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MOLECULARDIMENSIONSProviding you with the latestinnovations in protein crystallographyInsideLatest screensNew additivesCryo cocktailsIn situ platesSub µL DLSCrystal lead finderAcademic articlesCrystallization exercisesIntelligent solutions forprotein crystal growth

CONTENTS4 Crystal growth screens5 Soluble proteins5 Structure screen 16 Structure screen 26 Stura footprint screens7 Mini screen7 MultiXal screen8 The solubility tool kit8 Clear strategy screens I & II9 PACT premier9 JCSG plus13 Morpheus ®14 Protein complexes14 Macrosol15 Proplex15 MIDAS16 Nuclear receptors16 Nuclear receptor ligandbinding domain17 Membrane proteins17 MemStart18 MemSys18 MemPlus19 MemGold19 MemGold 222 Lipidic-sponge phase screen22 PGA screen23 MemMagic ® bicelle screen kit23 The Calixar Additive Kit24 MemAdvantage25 Nucleic acids25 Helix26 Optimization30 Custom screens31 Naomi’s nucleant31 MicroSeed Beads35 Trace Fluorescent labelling kits36 Additive screen36 Single reagents37 PEG precipitants38 Alternative precipitiants39 Jeffamines ®39 Volatiles – organics40 Non-volatiles – organics40 Polyamines41 Salts43 Other reagents43 Cryoprotectants44 Buffer stocks45 Special buffers45 Morpheus ® mixes46 Morpheus ® individualstock reagents48 MIDAS reagents49 MemAdvantage reagents51 The Really Useful Buffer kit51 Gelled Surface Kit51 Microbatch Oils60 Plates61 XRL 24–well plates61 24-well SBS plates61 Grease61 Sitting drop bridges62 Coverslips63 Counter diffusion66 High-throughput plates68 MRC Crystallization plates68 Triple drop plates68 MRC Maxi Optimization plates69 Microbatch plates69 In situ diffraction plates72 Hanging drop plates & seals72 Screw top hanging drop plate73 Laminex for LCP73 Sealing sheets and deep wellblocks76 Protein production76 Protein production80 Protein expression media80 Animal product free media81 Media optimization kit81 LB Broth82 Turbo Broth82 Superior Broth83 Power Broth83 Hyper Broth84 Glucose M9Y, & nutrient mix84 Augmedium85 LB Booster85 Atholate85 Perk expression rescue kit86 Expressmax86 Labelling media87 SelenoMethionine media87 SILAC media88 Cell culture products88 BRFF-BMZERO89 BRFF-EPM289 BRFF-HPC190 BRFF-P4-8F90 PET Cell dissociation formula91 Freezing media pair91 FNC Coating mix92 HEPES buffered saline92 DMEM / F12 serum free medium93 MDM serum free medium93 Bovine pituitary extract94 SFM screening kit94 Protein refolding reagents95 Quickfold refolding kit95 Detergent screening kit96 Cyclodextrin kit96 Dialysis & Accessories97 Gebaflex dialysis tubes97 Capillaries97 Paper wicks97 LCP mixing adaptor98 Cryocrystallography115 Books116 Tools118 Giftware102 LithoLoops102 Elliptical LithoLoops102 Mesh LithoLoops103 CryoMount Sets104 Magnetic CryoVials & CryoCaps105 DataMouse Pro105 Cryoprotectants109 CryoProtX110 Dewars110 Cryogenic shippers, & storagedewars112 EMBL/ESRF sample changer kits114 CryoTools & accessories116 Microtools117 Greasing119 Crystal structure glass blocks120Crystallography instrumentation120 Incubators120 Bench top incubators122 Floor standing vibration freeincubators124 Liquid handling124 Optimizer126 Imaging126 Crystal X2 automated lead finder128 X-taLight 200 automatedUV imaging system129 X-taLight 100 UV fluorescencelight source130 Dynamic light scattering130 SpectroSize 300 cuvette dls131 SpectroLight 600 sub µLscreening dls132 Pre crystallization screening133 In crystallization analysis134 Product Index145 How to orderNorth & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com 1

THE MEANING OF QUALITYQUALITY IS A WORD THAT IS OFTEN USED BUTRARELY CARRIES ANY REAL MEANINGISO 9001RegisteredQualityManagementEditorialWe read about “quality products” but find littlequalification of what is actually being described.Quality systems, and international qualityassurance standards are, however, verydifferent as they convey real and unequivocalmeaning. To achieve accreditation to aninternationally recognised standard requiresthe operation of a quality assurance system ofprocesses and procedures that are regularlyaudited internally and externally assessed inorder to maintain certification.Such internationally agreed standards are regularly updated andan accredited company must adapt its systems in order tocontinue to meet the published criteria. Molecular DimensionsLimited has achieved accreditation to ISO 9001: 2008 from theinternational assessment organisation NQA. This is part of the latestversion of the ISO 9000 standard and perhaps the biggest change fromprevious versions has been the emphasis on continual improvement. It isno longer good enough to continue to meet measurable performancelevels alone; we now have to demonstrate that we have systems that helpus to improve. Our measured trends also must show that we areconstantly improving the way we run our business.Our Quality System is not just a system of internal procedures but isdesigned to deliver real benefits to our customers. Our quality policystates that we will win your business by having the best reputation forreliable products, advice and service information, including delivery.The Molecular Dimensions Quality System is a set of processes that notonly establish conformance but, strive to continually improve oureffectiveness as a business. Our system has three main components:system processes and monitoring processes which ensure that the systemfunctions in a measurable way; but of most importance to you are thecustomer processes. We have standards for how we handle the completeproduct supply chain from how we answer customer inquiries to deliveringan order.Customer related processesEnquiriesOrder processingProductionFinal product monitoringGoods receivedStock controlPacking & deliveryNon-conforming productsNon-conforming serviceInside these processes there are procedures that are customer focused.For example our product standards are all set based on their expectedperformance in our customers’ laboratories. It is often said atconferences that there is a need for rationalisation of the number ofconditions used for screening for crystal growth based on what have beensuccessful. All of the Molecular Dimensions’ screens have been developedby our customers in just that way; as a result of selecting sets ofconditions that have worked particularly well in their experience. As aresult of this approach our product development programme is ‘trulycustomer focussed’.2 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

Welcome to the new Molecular Dimensionscatalogue. Looking back to fifteen years ago when we startedMolecular Dimensions it was our vision to offer crystallographersa choice of the very latest tools for their research and also bringnew methods to a broader audience all over the world. As witheverything we started small but rapidly became well recognisedin Europe and within two years had opened a subsidiary companyin the USA offering next day delivery from east to west coast.As the science of crystallography progressed we continued to fosterand maintain strong alliances with leading crystallographers to develop and commercialiseunique, proven ideas. Some of these have kindly contributed interesting and educationalarticles which can be found throughout this catalogue. In addition to this, MolecularDimensions formed partnerships with entrepreneurial companies with novel technologiesand ideas applicable to further bridging the gap between industrial and academic thinking.These collaborations have led to the introduction of crystallization methods for highefficiency screening, extended range crystallization space, regularly updated tools formembrane proteins, and stimulated high throughput methods for crystallization and datacollection, to name but a few. Most importantly continually delivering proven results hasgiven people the belief to try something new for the first time and also the ability to trust inour products for their day-to-day research.We know that major progress in scientific research is driven by the continuous developmentof new technology and methods, structural biology is no exception. Over time we have builta team of trained customer service staff and experienced scientists to understand both thework you do and also provide reliable technical and customer service information. Today weare proud to be supported by equally well qualified personnel throughout the globalcommunity of crystallography who are able to provide local service and accurate advice inmarkets such as India, the Far East and South America.In 2013 Molecular Dimensions continues its commitment to bring forward new and excitingideas with the introduction of a modern nucleic acid screen, a counter diffusion screen, thefirst membrane protein additive screen, calix(4)arene additives, fluorescent trace labelling,cryo-protection cocktails, high throughput plates for lipidic cubic phase, in situ screeningplates, and micro DLS directed crystal growth.We hope both these and the future developments we are committed to delivering willenable you to get the positive results you need.We are sure you will find the catalogue both interesting and informative. Good luck withyour research and we look forward to seeing you all again soon,Tony SavillPresident, Molecular DimensionsTony SavillPresident, Molecular DimensionsNorth & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com3

CRYSTAL GROWTH SCREENSMOLECULAR DIMENSIONS SUPPLIES A WIDE RANGE OF CRYSTALGROWTH SCREENS FROM CLASSIC SPARSE MATRICES TOMODERN HIGH EFFICIENCY PRODUCTS EXTENSIVELY COVERINGCRYSTALLIZATION SPACE FOR GLOBULAR PROTEINS, MEMBRANEPROTEINS AND NUCLEIC ACIDS.5 Soluble proteins5 Structure screen 16 Structure screen 26 Stura footprint screens7 Mini screen7 MultiXal screen8 The solubility tool kit8 Clear strategy screens I & II9 PACT premier9 JCSG plus14 Protein complexes14 Macrosol15 Proplex15 MIDAS16 Nuclear receptors16 Nuclear receptor ligandbinding domain17 Membrane proteins17 MemStart18 MemSys18 MemPlus13 Morpheus ® 19 MemGold19 MemGold 222 Lipidic-sponge phase screen22 PGA screen23 MemMagic ® bicelle screen kit23 The Calixar Additive Kit24 MemAdvantage25 Nucleic acids25 Helix4 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

SOLUBLE PROTEINSSOLUBLE PROTEINSSTRUCTURESCREEN 1STRUCTURE SCREEN 1 IS FORMULATED FOR THECRYSTALLIZATION OF PROTEINS, PEPTIDES, NUCLEICACIDS, & WATER SOLUBLE SMALL MOLECULES.This classic screen was originally published by Jancarik & Kimfrom conditions found to be successful in the crystallization ofbiological macromolecules.Features of Structure Screen 1:n The original sparse matrix screen 1 .n Enhanced buffer selection 2 .n Sparse matrix formula efficiently samples salts, polymers,organics & pH.n Proven effective with more than 1,000 biological macromolecules.n A complete kit designed to provide an effective and rapid screeningmethod for the crystallization of biological macromolecules.n A simple and practical way to find initial crystallization conditions.ORDER INFORMATIONDescription Pack CodeStructure Screen 1 50 x 10 mL MD1-01Structure Screen 1 cacodylate-free 50 x 10 mL MD1-01-CFThe Structure Screen Combination 100 x 10 mL MD1-03The Structure Screen Combination 100 x 10 mL MD1-03-CFcacodylate-freeStructure Screen I + II HT-96 96 x 1 mL MD1-30Structure Screen I + II HT-96 96 x 1 mL MD1-30-CFcacodylate-freeStructure Screen 1 Single Reagent 100 mL MDSR-01Structure Screen 1 Single Reagent 250 mL MDSR-01-250Structure Screen 1 + 2 HT-96 100 mL MDSR-30Single ReagentReferences1. Jancarik, J. & Kim, S.H. J. Appl. Cryst. (1991), 24, 409-411.2. Wooh et al Acta Cryst. (2003) D59, 769 – 772.North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com5Crystal Growth Screens

Crystal Growth ScreensSOLUBLE PROTEINSSTRUCTURESCREEN 2STRUCTURE SCREEN 2 IS FORMULATED FOR THECRYSTALLIZATION OF PROTEINS, PEPTIDES, NUCLEICACIDS, & WATER SOLUBLE SMALL MOLECULES.SOLUBLE PROTEINSSTURA FOOTPRINTSCREENSTHE FOOTPRINT SCREENS ARE BASED ON THECONCEPT OF SCREENING THE PROTEIN PRECIPITANTSOLUBILITY CURVE: A PROTEIN SOLUBILITY SCREEN.A classic extension to the screen originally published by Jancarik &Kim 1 from conditions found to be successful in the crystallization ofbiological macromolecules.The screens are presented as two sets of 24 conditions at acidic,neutral and basic pH's for polyethylene glycols and salts.Features of Structure Screen 2:n An extension 2 to the original 1 sparse matrix screen with novelprecipitants and combinations.n Structure Screen 2 contains 50 different and unique reagentcombinations.n Sparse matrix formula efficiently samples salts, polymers, organics& pH.n Proven effective with more than 1,000 biological macromolecules.n A complete kit designed to provide an effective and rapid screeningmethod for the crystallization of biological macromolecules.n A simple and practical way to find initial crystallization conditions.Features of Footprint Screening:n Screen the protein precipitant solubility curve rather than acrystallization trial 1 .n Test the relative protein solubility with precipitants that have beenused successfully in the crystallization of many proteins 2 .n Once initial crystals or crystalline aggregates are obtained from theinitial screening, streak seeding is recommended to determine theranges of conditions under which crystal growth can proceed 3 .ORDER INFORMATIONDescription Pack CodeStructure Screen 2 50 x 10 mL MD1-02Structure Screen I + II HT-96 96 x 1 mL MD1-30Structure Screen I + II HT-96 96 x 1 mL MD1-30-CFcacodylate-freeThe Structure Screen Combination 100 x 10 mL MD1-03The Structure Screen Combination 100 x 10 mL MD1-03-CFcacodylate-freeStructure Screen 2 Single Reagent 100 mL MDSR-02Structure Screen 1 + 2 HT-96 100 mL MDSR-30Single ReagentReferences1. Jancarik, J & Kim, S.H., J. Appl. Cryst. (1991) 24, 409-411.2. Cudney, R., Patel, S., Weisgraber, K., Newhouse, Y., and McPherson, A., Acta Cryst. (1994) D50, 414-423.ORDER INFORMATIONDescription Pack CodeStura Footprint Screens 48 x 10 mL MD1-20Stura Footprint Screen cacodylate-free 48 x 10 mLMD1-20-CFThe Stura Footprint Combination 96 x 10 mL MD1-05The Stura Footprint Combination 48 x 10 mL MD1-05-CFcacodylate-freeStura Footprint Combination HT-96 96 x 1 mL MD1-43Stura Footprint Combination HT-96 96 x 1 mL MD1-43-CFcacodylate-freeStura Footprint Screen Single Reagent 100 mL MDSR-20-Stura MacroSol HT-96 Single Reagent 100 mL MDSR-43-*Combination includes Stura Footprint Screens and Macrosol (p14).References1. Stura E.A., Nemerow G.R., Wilson I.A, (1992),. Journal of Crystal Growth 122, 273-285.2. Stura E.A. (1999) Strategy 3: Reverse Screening. In "Crystallization of Proteins: Techniques, Strategiesand Tips. A laboratory manual" (Bergfors T. ed.) International University Line pp113-124.3. Stura E.A, Satterthwait A.C., Calvo J.C., Kaslow D.C., Wilson I.A. (1994) Reverse Screening. Acta Cryst.D50: 448-455.6 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

SOLUBLE PROTEINSMINI SCREENA MINIMISED SPARSE MATRIX SCREEN BASED ON THECLASSIC JANCARIK AND KIM SCREEN 1 BUT WITH MINIMALREDUNDANCY.SOLUBLE PROTEINSMULTIXTAL CRYSTALLIZATION SCREENTHE FLEXIBLE CRYSTALLIZATION SCREENThe kit contains a 24 condition, sparse matrix screen.Features of Mini Screen:n 24 conditions optimised for maximal probability of successfullyobtaining a crystal with minimal redundancy 2 .n Particularly suitable as a cost effective start to crystallizationexperiments or where material is limited.n In a structural proteomics context, the kit works as an efficient,optimized screen that will give maximum samples for structuresolution while minimising the amount of time and material wastedon unnecessary experiments.ORDER INFORMATIONDescription Pack CodeMini Screen 24 x 10 mL MD1-09Mini Screen cacodylate-free 24 x 10 mL MD1-09-CFMini Screen Single Reagent 100 mL MDSR-09-References1. Jancarik J, Kim SH., (1991) , J. Appl. Cryst. 24, 409-411.2. Kimber M.S. et al., (2003), Proteins: Structure, Function, and Genetics 51, 562-568.3D STRUCTURE SCREEN3D STRUCTURE SCREEN IS FORMULATED FOR GROWINGPROTEIN CRYSTALS IN THE METASTABLE ZONE USING THEVAPOUR DIFFUSION TECHNIQUE.This method is presented in the 3D Structure Screen as anoptimized sparse matrix crystal growth screen which allows the24 “nucleation” conditions of the Mini Screen and 24 “backed-off”conditions for crystal growth to be tried. Formatted for HT blocksas the Heavy plus Light Twin Pack HT-96.Features of 3D Structure Screen:n Separate nucleation from crystal growth.n A simple method for growing crystals in the metastable zone.n Obtain much larger and better diffracting crystals compared toconventional methods.ORDER INFORMATIONDescription Pack Code3D Structure Screen 48 x 10 mL MD1-133D Structure Screen cacodylate-free 48 x 10 mL MD1-13-CFHeavy + Light Twin pack HT-96 2 x 48 x 1 mL MD1-35Heavy + Light Twin pack HT-96 2 x 48 x 1 mL MD1-35-CFcacodylate-free3D Structure Screen Single Reagent 100 mL MDSR-13-Heavy & Light HT-96 Single Reagent 100 mL MDSR-35-This product is manufactured under an exclusive licence from Imperial College of Science,Technology and Medicine, London U.K.See page 57 for hints on screening using this screen.A great addition to any crystallization lab – a 48 condition sparsematrix crystallization screen offering multiple uses forcrystallization. For soluble and membrane proteins.Use it as a high PEG screen for vapour diffusion as well as incounter-diffusion experiments and seeding experiments. Containshigher PEG concentrations than found in traditional standardscreens; offering greater flexibility in its usage.Features to come:n High PEG concentration screen.n Suitable for soluble and membrane proteins.n Ideal for counter-diffusion experiments – especially compatible withCrystalHarp .n Use in seeding experiments.n Available as 10 mL, HT or prefilled microplate (FX) format.ORDER INFORMATIONDescription Pack CodeMultiXtal 48 x 10 mL MD1-65MultiXtal HT-96 (48 x 1 mL) x 2 MD1-66MultiXtal FX(48 x 0.1 mL) x 2 MD1-67(pre-filled conical bottom microplate)MultiXtal Single Reagent 100 mL MDSR-65MultiXtal HT-96 Single Reagent 100 mL MDSR-66MultiXtal FX Single Reagent 100 mL MDSR-67See also Crystal Harp (p65).North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com7Crystal Growth Screens

Crystal Growth ScreensSOLUBLE PROTEINSTHE SOLUBILITYTOOL KIT A SYSTEMATIC ALTERNATIVE TO SPARSEMATRICES TO DETERMINE THE SOLUBILITY ANDCRYSTALLIZATION POTENTIAL OF PROTEIN SALTS 1 - 4.SOLUBLE PROTEINSCLEAR STRATEGYSCREEN I / CLEARSTRATEGY SCREEN IITWO COMPLIMENTARY 6 X 4 MATRIX SCREENSDEVELOPED BY DR. A. M. BRZOZOWSKI AND J.WALTON 1 FROM THE LABORATORY OF STRUCTURALBIOLOGY AT YORK UNIVERSITY.Each kit contains 10 mL of 24 reagents buffered at pH 4.5, 24reagents buffered at pH 9.0 (box 1) , 18 unbuffered polyethylene glycols,6 unbuffered MPD’s, 21 salt concentrates and buffer concentrates(box 2)from pH 5.5 – 9.5.Originally these screens were tested on a number of proteins, whichhad not been crystallized previously and yielded diffraction qualitycrystals. Now these are amongst our most popular products.Each screen contains 24 stock solutions and the following (1M)buffers: Sodium Acetate, pH 4.5; Sodium Acetate, pH 5.5; SodiumCacodylate, pH 6.5; Tris, pH 7.5; and Tris, pH 8.5.Features of The Solubility Tool Kit:n Provides the best starting conditions for the right buffer, pH and salt.n Rapidly identifies likely crystallization reagents and their suitablerange of concentration.n Locate the nucleation zone of a protein and prepare phasediagrams.n Can be performed with batch, vapour diffusion or dialysis technique.n Provides detailed information on the solubility profile of a protein.ORDER INFORMATIONDescription Pack CodeThe Solubility Tool Kit (Box 1 + 2) 98 x 10 mL MD1-17Solubility Screen Box 1 48 x 10 mL MD1-18Precipitant Screen Box 2 50 x 10 mL MD1-19The Solubility Tool Kit Single Reagent 100 mL MDSR-17-References1. Riès-Kautt M, Ducruix A,. Methods Enzymol 1997, 276:23-59.2. Riès-Kautt M, Ducruix A, In "Crystallization of Nucleic acids and proteins: A practical approach". (Ducruix Aand Giegé R. ed.) IRL/Oxford Press.3. Riès-Kautt M Strategy 2, In "Protein Crystallization Techniques, Strategies, and Tips" (Bergfors T. ed) IULBiotechnology Series International University Line, La Jolla, California.4. Vaney M. C. et al, Acta Cryst D (2001), D57, 929-940.Features of Clear Strategy Screens:n Limit number of trials.n Aid rational design of subsequent trials.n User defined pH.n Use protein information.n Maintain ‘folding homogeneity’ of protein.n Interchangeable components.n Potential anomalous scattering centres 2 .ORDER INFORMATIONDescription Pack CodeClear Strategy Screen I 24 x 10 mL* MD1-14Clear Strategy Screen I cacodylate-free 24 x 10 mL*MD1-14-CFClear Strategy Screen II 24 x 10 mL* MD1-15Clear Strategy Screen II cacodylate-free 24 x 10 mL*MD1-15-CFThe Clear Strategy Screen Combination 48 x 1 mL* MD1-16The Clear Strategy Screen 48 x 10 mL* MD1-16-CFCombination- cacodylate-freeClear Strategy Screen Combination 240 x 10 mL MD1-16LMBpH PremixedClear Strategy Screen I HT-96 96 x 1 mL MD1-31Clear Strategy Screen I HT-96 96 x 1 mL MD1-31-CFcacodylate-freeClear Strategy Screen II HT-96 96 x 1 mL MD1-32Clear Strategy Screen II HT-96 96 x 1 mL MD1-32-CFcacodylate-freeClear Strategy Screen 1 Single Reagent 100 mL MDSR-14-Clear Strategy Screen 11 Single Reagent 100 mL MDSR-15-Clear Strategy Screen 1 HT-96 Single Reagent 100 mL MDSR-31-Clear Strategy Screen 2 HT-96 Single Reagent 100 mL MDSR-32-* Plus buffers.These products are manufactured under an exclusive license from theUniversity of York, UK.References1. Brzozowski A. M. and Walton J., (2001) J. Appl. Cryst. 34, 97-101.2. Dauter Z., Dauter M. & Rajashankar K. R. (2000), Acta Cryst. D56, 232-237.3. Selmer et al, (2006), Science 313, 1935-1442.8 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

SOLUBLE PROTEINSSOLUBLE PROTEINSPACT PREMIER PACT PREMIER IS A pH, ANION, CATION CRYSTALLIZATIONTRIAL DEVISED TO TEST PH WITHIN A PEG/ION SCREENENVIRONMENT. THIS IS ONE OF THE MOST EFFECTIVESYSTEMATIC SCREENS AVAILABLE TO DATE.Building on the success of PEG/Ion screening, this PEG/ION/pHscreen has been developed to systematically test the effect of pH,anions and cations, using PEG as the precipitant. This screen wasoriginally implemented very successfully at the Netherlands CancerInstitute (NKI), and at the Oxford Protein Production Facility(OXPPF). The kit contains 96 reagents arranged as a cation/PEGscreen, an anion/PEG screen and a pH/PEG screen.Features of PACT Premier:n A modern, comprehensive PEG/ion screen.n This 96-well screen is really 3 screens:1. 24-well pH/PEG screen.2. 24-well cation/PEG screen.3. 48-well anion/PEG screen.ORDER INFORMATIONDescription Pack CodePACT premier (10 mL kit) 96 x 10 mL MD1-29PACT premier cacodylate-free 96 x 10 mL MD1-29-CFPACT premier HT-96 96 x 1 mL MD1-36PACT premier HT-96 cacodylate-free 96 x 1 mL MD1-36-CFPACT premier HT-96 Green Screen 96 x 1 mL MD1-52PACT Green Screen (10 mL) 96 x 10 mL MD1-55PACT premier Screen Single Reagent 100 mL MDSR-29-PACT premier Screen HT-96Single Reagent 100 mL 100 mL MDSR-36-PACT premier HT-96 Green ScreenSingle Reagent 100 mL MDSR-52-PACT premier Green Screen (10mL)Single Reagent 100 mL MDSR-55-Super2 Combo Value Pack(JCSG+ & PACT premier)2 x 96 x 10 mL MD1-75The screen was developed by Janet Newman, and was tested inthe laboratory of Anastassis Perrakis at the NKI as part of theSPINE programme, and is manufactured under license byMolecular Dimensions.References1. Newman et al, (2005), Acta Cryst. D61, 1426JCSG-PLUS SCREENAN OPTIMIZED SPARSE MATRIX SCREEN OF CLASSICAND MODERN CONDITIONS DEVISED AT THE JOINTCENTRE FOR STRUCTURAL GENOMICS 1 ANDDEVELOPED FURTHER BY NEWMAN AND PERRAKIS 2AT THE NETHERLANDS CANCER INSTITUTE.The kit contains a 96 condition, sparse matrix screen.Features of JCSG-Plus:n Optimized sparse matrix screen.n Reduced redundancy.n Screens classic PEG and salt conditions.n Access more areas of crystallization space.n Neutralised organic acids:Formate, acetate, citrate, succinate, malate, malonate 3 .n More organic and polyalcohol conditions.n Precipitant synergy.n Wide pH range 4.0 – 10.0.ORDER INFORMATIONDescription Pack CodeJCSG plus 96 x 10 mL MD1-37JCSG plus cacodylate-free 96 x 10 mL MD1-37-CFJCSG plus HT-96 96 x 1 mL MD1-40JCSG plus HT-96 cacodylate-free 96 x 1 mL MD1-40-CFJCSG plus HT-96 Green Screen 96 x 1 mL MD1-53JCSG Plus Green Screen (10 mL) 96 x 10 mL MD1-56Super2 Combo Value Pack2 x 96 x 10 mL MD1-75(JCSG+ & PACT)JCSG plus Screen Single Reagent 100 mL MDSR-37-JCSG plus HT-96 Single Reagent 100 mL MDSR-40-JCSG plus HT-96 Green Screen 100 mL MDSR-53-Single ReagentJCSG Green Screen (10 mL) 100 x 10 mL MDSR-56-Single ReagentReferences1. Page et al (2003), Acta Cryst. D59, 1028-1037.2. Newman et al (2005), Acta Cryst. D61, 1426-1431.3. McPherson et al (2001), Protein Science 10, 418422.North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com9Crystal Growth Screens

TOWARDS HIGHEFFICIENCY SCREENING“…conditions most essential for promoting crystal formation…”Page et al (2003) Acta Cryst. D59, 1028EditorialSPARSE MATRICESThe earliest sparse matrix screens were based onpreviously successful crystallization conditions.Since most researchers used these screens, thesame sub-sets of conditions were used repeatedly.As a result of the published results of data-miningexercises from a number of structural genomicsinitiatives more efficient screens have been devised.The Joint Centre for Structural Genomics (JCSG) analysed thecrystallization of over 500 different proteins against availablesparse matrices. Using a novel algorithm, members of the JCSGidentified a core screen (JCSG) developed when data miningrevealed massive redundancy between clusters of conditions particularlywhere high molecular weight PEGs were used as precipitants (1).The second issue to come to light was that even extensive suites of sparsematrix screens represent incomplete coverage of crystallisation space –480 conditions failed to crystallise 15% of the target proteins.The JCSG-plus screen (p9) developed at the Netherlands Cancer Instituteby Janet Newman during the SPINE project supplements the JCSG coreconditions to provide a more complete coverage of crystallisation space(2), by in-filling the pH profile, introducing neutralised organic acids asthe precipitant (3) and expanding the range of organic and polyalcoholconditions to create precipitant synergy.Rational approachesThere are a few screens that try to test crystallization space in a morerational manner – for example, the Clear Strategy Screen (4) (p8), and theSolubility Tool Kit (5) (p8). The PEG/Ion screen (6) has been reported tobe very successful in producing crystal hits, with the caveat that many ofthe hits are redundant.With this in mind, the PEG/ION/pH screen (PACT-premier) (p9) wasdeveloped (2) to systematically and additionally test the effect of pH, aswell as anions and cations, using PEG as the precipitant. This screen hasbeen implemented very successfully at the Netherlands Cancer Institute(NKI), and at the Oxford Protein Production Facility (OXPPF).The JCSG-plus sparse matrix screen is highly effective when usedalongside a systematic screen such as PACT-premier. The two screensprovide a thorough exploration of crystallization conditions and theunique design of PACT-premier facilitates rational interpretation ofresults from both itself and JCSG-plus assisting the design of subsequentexperiments.References1. Page et al (2003). Shotgun crystallization strategy for structural genomics: an optimizedtwo-tiered crystallization screen against the Thermotoga maritima proteome.Acta Cryst. D59, 1028-10372. Newman et al (2005). Towards rationalization of crystallization screening for smalltomedium-sized academic laboratories: the PACT/JCSG+ strategy.Acta Cryst. D61, 1426-14313. McPherson et al (2001). A comparison of salts for the crystallisation of macromolecules,Protein Science 10, 4184224. Brozowski & Walton (2001) J. Appl. Cryst. 34, 97-1015. Ries-Kaut in Protein Crystallization techniques,. Strategies and Tips (Bergfors T Ed.)IUL Series.6. A comparison of salts for the crystallization of macromolecules.Alexander McPherson. Protein Science (2001), 10:418-422.10 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

Dr Rebecca PageDr Janet NewmanNorth & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com11

EXPLORING MORECRYSTALLIZATIONSPACEEditorialDr Clemens Grimm (Würzburg).Out of 8289 entries scanned in the PDB, almost half of thecrystallization conditions contained a PEG component andmost commercial screens contain PEGS. However, thesuccess rate of PEGS might be biased due to theirwidespread dominance in crystallization screens. There are manyalternatives to PEGs and a variety have recently been described asbeing useful for macromolecular crystallogenesis, but had onlysporadically been introduced into standard crystallization screens.To close this gap, Clemens Grimm et al (1) at Würzburg University inGermany, have devised a protein crystallization screen (MIDAS) thatsystematically searches for crystallization conditions with alternativepolymeric precipitants.MIDAS covers a relatively narrow range of pH and salt concentrationscentred on physiological values to increase its suitability for sensitivemacromolecular complexes, while every condition contains at least onealternative polymeric precipitant.MIDAS has been designed to complement PEG- and salt-basedscreens, and is ideal for protein, protein/protein complexes, proteinnucleicacid complexes and sensitive macromolecular complexes.Dr AM Brzozowski (York)FOR DECADES PEGS OR THEIRMONOMETHYL ETHERS (PEG MMES),HAVE DOMINATED CRYSTALLIZATIONSCREENSAn alternative new systematic screen from the York Structural BiologyLaboratory (YSBL) based on the poly-γ-glutamic acid (PGA) polymerhas also been reported (2). This PGA Screen (p22) is suitable for bothglobular and membrane protein crystallization. This novel precipitantallows totally new crystallization space to be explored.PGAs present at least two new aspects in protein crystallization; theyextend the range of existing PEG-based polymers into: new-chemicalpolymers that exploit poly-amino acids and broaden the range ofmolecular weight of polymer precipitants into regions over 1MDa.12 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

The high nucleation-precipitation potential of PGAs enables theiruse at very low concentrations and in combination with classicalprecipitants, thus scaling down the amount of precipitantnecessary for crystal appearance and growth. This feature of PGAsmakes them especially useful in applications for labile, easilyprecipitating proteins.Although they can be employed for all type/classes of proteins,current experience resulting from work in the YSBL suggests thatPGA should be especially effective for crystallization of membraneproteins. Therefore, the PGA-based screens are recommended astargeted screens with membrane proteins as the main/primarysubjects of their applicability.Morpheus (right) aims to unlock novel chemical space previouslyinaccessible using conventional screens by incorporating a rangeof low molecular weight ligands found ordered in more than 33,000deposited structures. This 96 condition 3D crystallization screenwas developed and utilized in the world renowned Laboratory ofMolecular Biology, Cambridge, UK.Morpheus is based on extensive data mining of the PDB. The aim isto explore different chemical space than that achieved withconventional screening. Morpheus incorporates 49 low molecularweight components. They are PDB ligands sharing four maincharacteristics; they are small (the largest being HEPES MW 238.30g/mol and the smallest a lithium ion MW 6.94 g/mol), stable,inexpensive and are associated with at least five unrelated PDBstructures. For instance, the two enantiomers of tartaric acid(PDB ID: TAR and TLA) are found ordered in 113 structures.References1. Grimm, C., Chari, A., Reuter, K. & Fischer, U. (2010). Acta Cryst. D66, 685-697.2. TC Hu, J Korczynska, DK Smith, AM Brzozowski - Acta Crystallographica Section D: BiologicalCrystallography, 2008. D64, 957-9633. Gorrec F. (2009) J. Appl. Cryst. 42, 1035-1042SOLUBLE PROTEINSMORPHEUS ®A 96 CONDITION PROTEIN CRYSTALLIZATION SCREEN WITHORIGINAL CHEMISTRY INCORPORATING A RANGE OF LOWMOLECULAR WEIGHT LIGANDS FOUND ORDERED IN MORETHAN 33000 PDB STRUCTURES.Developed and utilized in the world renowned Laboratory ofMolecular Biology, (Cambridge, UK) - Morpheus ® carries on thetradition of being original and dynamic. This unique screen aims tounlock novel chemical space previously inaccessible usingconventional screens.The ligands most frequently reported in PDB structures are groupedinto eight mixes of additives based on chemical class (e.g. alcohols,carboxylic acids, etc). Several of these top PDB ligands are“biological buffers” like HEPES and have been used to build threebuffer systems. Nine precipitants are grouped into four mixes thatcontain at least a PEG (Polyethylene glycol) and a different type ofprecipitant that is also a cryo-protectant (e.g. Glycerol).The kit is presented as a 96 condition screen.Features of Morpheus ® :n Simple and effective 3D grid design covering a range of pH, PEGs andsalt additives.n 49 low molecular weight ligands promote both initial crystal formationand lattice stability.n Reduced crystal "stress" - all conditions are cryo-protected.n Derived from extensive data mining of over 33,000 PDB entries.n Available in both 10 mL tube and 96 condition deep well blockformats.ORDER INFORMATIONDescription Pack CodeMorpheus 96 x 10 mL MD1-46Morpheus HT-96 96 x 1 mL MD1-47Morpheus HT-96 Green Screen 96 x 1 mL MD1-47-GreenMorpheus Additive OptiMax Kit (10 mL) 43 x 10 mL MD1-58Power Combo Value Pack2 x 96 x 10 mL MD1-76(Morpheus & MIDAS)Morpheus Screen Single Reagent 100 mL MDSR-46-Morpheus HT-96 Single Reagent 100 mL MDSR-47-Crystal Growth ScreensThis product has been designed and developed by Fabrice Gorrec, in collaboration with the scientists atthe Medical Research Council Laboratory of Molecular Biology (LMB) at Cambridge and is manufacturedexclusively under license by Molecular Dimensions Limited.Fabrice Gorrec receives the MRC CEO's Award from John Savill for innovations in the field ofmacromolecular crystallization (including the Morpheus screen).ReferencesGorrec F. (2009). J. Appl. Cryst. 42, 1035-1042North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com13

Crystal Growth ScreensPROTEIN COMPLEXESPROTEIN COMPLEXESMACROSOL THIS FOOTPRINT SCREEN FOR THE CRYSTALLIZATION OFPROTEIN COMPLEXES AND OTHER SAMPLES WITHHETEROGENEOUS SOLUBILITY, SCREENS THE PROTEINPRECIPITANT SOLUBILITY CURVE RATHER THAN SETTINGUP A RANDOMIZED CRYSTALLIZATION TRIAL. 1-4The screens are presented as two sets of 24 conditions at acidic,neutral and basic pH's for polyethylene glycols and salts.Features of Macrosol:n Screens the protein precipitant solubility curve rather than acrystallization trial.n Satisfies dual requirements:– Maintain protein-protein interactions.– Reduce solubility of complex.n Test the relative protein solubility with precipitants that have beenused successfully in the crystallization of many proteins.n Once initial crystals or crystalline aggregates are obtained from theinitial screening, streak seeding is recommended to determine theranges of conditions under which crystal growth can proceed.ORDER INFORMATIONDescription Pack CodeMacroSol 48 x 10 mL MD1-22MacroSol cacodylate-free 48 x 10 mL MD1-22-CFMacroSol Screen Single Reagent 100 mL MDSR-22-The Stura Footprint Combination 96 x 10 mL MD1-05The Stura Footprint Combination 48 x 10 mL MD1-05-CFcacodylate-freeStura Footprint Combination HT-96 96 x 1 mL MD1-43Stura Footprint Combination HT-96 96 x 1 mL MD1-43-CFcacodylate-freeReferences1. Stura E.A., Nemerow G.R., Wilson I.A (1992), Journal of Crystal Growth 122, 273-2852. Stura E.A. (1999) Strategy 3: Reverse Screening. In "Crystallization of Proteins: Techniques, Strategies andTips. A laboratory manual" (Bergfors T. ed.) International University Line pp113-124.3. Stura E.A, Satterthwait, A.C, Calvo, J.C, Kaslow, D.C, Wilson, I.A. (1994), Acta Cryst. D50, 448-455.4. Stura E.A, "EMBO Workshop on the crystallization of Macromolecular Complexes", Grenoble 8-13 April 200114 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

PROTEIN COMPLEXESPROPLEX PROPLEX IS FORMULATED FOR THECRYSTALLIZATION OF PROTEIN COMPLEXES.PROTEIN COMPLEXESMIDAS MIDAS IS A MODERN INTELLIGENT DYNAMIC ALTERNATIVESCREEN WITH 96 REVOLUTIONARY CRYSTALLIZATIONSCREENING CONDITIONS BASED ON ALTERNATIVEPOLYMERIC PRECIPITANTS DEVELOPED AND TESTED INTHE LABORATORY OF DR. CLEMENS GRIMM AT UNIVERSITYOF WÜRZBURG, GERMANY.Crystal Growth ScreensDeveloped and tested by Sergei Radaev et al 1,2 , at the NIH NationalInstitute of Allergy and Infectious Diseases.The kit contains a 96 condition, targeted sparse matrix screen.MIDAS is a revolutionary crystallization screen that has moved awayfrom the reliance on polyethylene glycols (PEGs) as the mainprecipitant (only 3 conditions in MIDAS contain a PEG).Features of ProPlex:n Targeted sparse matrix screen.n Based on results of Protein Complex Crystallization Database.n Satisfies dual requirements:– Maintain protein-protein interactions.– Reduce solubility of complex.n Medium and High MW PEGs.n Lower PEG concentrations.n Fewer organic precipitants.n Neutralised organic acids.n pH range from 4.0 – 8.5.Features of MIDAS:n Ideal for both protein, protein/protein complexes, protein / nucleicacid complexes, and sensitive macromolecular complexes.n Narrow range of pH and salt concentrations centered onphysiological values.n Every condition contains at least one alternative polymericprecipitant.n Compatible with liquid handling robots.ORDER INFORMATIONDescription Pack CodeProPlex 96 x 10 mL MD1-38ProPlex cacodylate-free 96 x 10 mL MD1-38-CFProPlex HT-96 96 x 1 mL MD1-42ProPlex HT-96 cacodylate-free 96 x 1 mL MD1-42-CFProPlex Screen Single Reagent 100 mL MDSR-38-ProPlex HT-96 Single Reagent 100 mL MDSR-42-ORDER INFORMATIONDescription Pack CodeMIDAS 96 x 10 mL MD1-59Power Combo Value Pack2 x 96 x 10 mL MD1-76(Morpheus & MIDAS)MIDAS HT-96 96 x 1 mL MD1-60MIDAS OptiMax Kit 27 x 10 mL MD1-62MIDAS Single Reagent 100 mL MDSR-59MIDAS HT-96 Single Reagent 100 mL MDSR-60References1. Radaev S., Li S. and Sun P. D. (2006), Acta Cryst. D62, pp 605-612.2. Radaev S., Li S. and Sun P. D. (2002), J. Appl. Cryst. 35, 674-6763. Dafforn (2007) So how do you know you have a macromolecular complex? Acta Cryst. D63, 17-25.4. Crystallisation of Nucleic Acids and Proteins, Edited by A. Ducruix and R. Giegé, The Practical ApproachSeries, Oxford Univ. Press, 1992.5. Protein Crystallization Techniques Strategies & Tips, (Bergfors T. ed.) IUL 1999.MIDAS is manufactured and distributed under an exclusive license with Dr. C. Grimm and Prof. Dr. U. Fischer.References1. Grimm, C., Chari, A., Reuter, K. & Fischer, U. (2010). Acta Cryst. D66, 685-697North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com15

Crystal Growth ScreensNUCLEAR RECEPTORSNUCLEAR RECEPTORSNUCLEAR RECEPTOR -LIGAND BINDINGDOMAIN SCREENSA RATIONAL APPROACH TO SCREENING INITIALCRYSTALLIZATION CONDITIONS OF NUCLEARRECEPTOR LIGAND BINDING DOMAINS. 1The NR-LBD Screen contains 48 conditions divided into two parts:24 conditions using various PEGs as the precipitant and 24conditions using various carboxylic acids as the precipitant.The extension contains a set of 48 unique reagents which not onlyallows the user to reproduce previously obtained results but alsoto crystallize new ligand binding domains.Features of the Nuclear Receptor -Ligand Binding Domain Screens:n Nuclear receptors usually crystallize in the presence of two reagentfamilies: polyethylene glycol and carboxylic acids.n The pH range for crystallization is 6.0 to 9.0.n Includes PEGs 4000, 6000 and 8000.n Includes sodium formate, sodium citrate, sodium acetate carboxylicacids.n Ionic strength controlled with sodium chloride and ammoniumacetate.n pH range is between 6.5 and 8.5 using Bis Tris (pH 6.5), PIPES(pH 7.0), Na HEPES (pH 7.5) and Tris (pH 8.0 and 8.5).ORDER INFORMATIONDescription Pack CodeNR-LBD Screen 48 x 10 mL MD1-24NR-LBD Extension Screen 48 x 10 mL MD1-26Nuclear Receptor Combination 96 x 10 mL MD1-27NR-LBD + NR-LBD Extension HT-96 96 x 1 mL MD1-34NR-LBD Screen Single Reagent 100 mL MDSR-24-NR-LBD Extension ScreenSingle Reagent 100 mL MDSR-26-NR-LBD + NR-LBD ExtensionHT-96 Single Reagent 100 mL MDSR-34-The NR-LBD screens were developed by Denis Zeyer, Sylvie Duclaud,Dino Moras and Jean-Paul Renaud at The Institute of Genetics andMolecular and Cell Biology, CNRS, UMR 7104, Illkirch, France and ismanufactured under an exclusive license from CNRS, France.References1. Zeyer D., Duclaud S., Moras D. Renaud J-P., Billas I. M. et al. (2001) J. Biol. Chem. 276, 7465-7474.16 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

MEMBRANE PROTEINSMEMBRANE PROTEINSMEMSTART A STARTING POINT FOR SCREENING AND OPTIMIZINGCRYSTALLIZATION CONDITIONS FOR ALPHA TYPETRANSMEMBRANE PROTEINS USING VAPOURDIFFUSION METHODS. 1A sparse matrix of 48 conditions allowing the pH range, precipitantsand salts used in membrane protein crystallization to be screenedwith detergent- containing protein drops.Molecular Dimensions acknowledges the work of Prof. S. Iwata,Dr. M. Iwata and Dr. J. Abramson in designing this product.Features of MemStart:n Based on the reagents typically used in the laboratory of Prof. S.Iwata.n Optimized to span 33 reported successful crystallization conditionsof membrane proteins for which atomic resolution structures havebeen determined.n Includes the pH, precipitant concentration and type, and salts foundto be successful.Crystal Growth ScreensORDER INFORMATIONDescription Pack CodeMemStart Kit 48 x 10 mL MD1-21MemStart Screen Single Reagent 100 mL MDSR-21-The Membrane Protein Combination 96 x 10 mL MD1-04MemStart + MemSys HT-96 96 x 1 mL MD1-33MemStart + MemSys HT-96 100 mL MDSR-33-Single ReagentThis product is manufactured under an exclusive licence from ImperialCollege of Science, Technology & Medicine, London, UK.References1. Methods and Results in the crystallization of Membrane proteins (2003) Ed. Iwata S. (InternationalUniversity Line)North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com17

Crystal Growth ScreensMEMBRANE PROTEINSMEMSYS A SYSTEMATIC SCREEN SPANNING THE KEY VALUESOF pH, PRECIPITANT TYPE/ CONCENTRATION, ANDSALTS FOR ALPHA TYPE TRANSMEMBRANEPROTEINS USING VAPOUR DIFFUSION METHODS. 1MEMBRANE PROTEINSMEMPLUS A FURTHER 48 CONDITIONS TARGETED AT MEMBRANEPROTEINS.48 conditions allowing the pH range, precipitants and salts used inmembrane protein crystallization to be screened with detergentcontaining protein drops. The reagents can be easily arranged insystematic array to facilitate the interpretation of results and thedesign of further optimization experiments.A useful addition to MemGold. This screen further targets β-barreltype membane proteins – including porins and outer membraneactive transporters. Crystallization conditions were data minedfrom the PDB.Features of MemSys:n A systematic approach to screening for initial crystallizationconditions for membrane proteins using vapour diffusion methods.n Membrane protein solubility is pushed to the limit to provide moreinformation than previous sparse matrix type screens.n Includes the pH, precipitant concentration and type, and salts foundto be successful.n Primarily designed for alpha type transmembrane proteins, but hasalso been successfully applied to beta type outer membraneproteins.Developed by members of the So Iwata Lab, Imperial College, London.Features of MemPlus:n Based on successful crystallization conditions data mined fromthe Protein Data Bank.n Contains 48 conditions covering a range of pH, PEGs andsalt additives.ORDER INFORMATIONDescription Pack CodeMemSys 48 x 10 mL MD1-25The Membrane Protein Combination 96 x 10 mL MD1-04MemSys Single Reagent 100 mL MDSR-25-MemStart + MemSys HT-96 96 x 1 mL MD1-33MemStart + MemSys HT-96 100 mL MDSR-33-Single ReagentThis product is manufactured under an exclusive licence from ImperialCollege of Science, Technology & Medicine, London, UK.References1. Methods and Results in the crystallization of Membrane proteins (2003) Ed. Iwata S. (InternationalUniversity Line)ORDER INFORMATIONDescription Pack CodeMemPlus 48 x 10 mL MD1-44MemPlus cacodylate-free 48 x 10 mL MD1-44-CFMemPlus HT-96 96 x 1 mL MD1-45MemPlus HT-96 cacodylate-free 96 x 1 mL MD1-45-CFMemPlus Single Reagent 100 mL MDSR-44-MemPlus HT-96 Single Reagent 100 mL MDSR-45-This product is manufactured under an exclusive licence from ImperialCollege of Science, Technology & Medicine, London, UK.References1. Newstead, Simon; Hobbs, Jeanette; Jordan, Davina; Carpenter, Elisabeth; Iwata, So, "Insights into outermembrane protein crystallisation." Molecular Membrane Biology (2008). Dec, 25(8): 631-638.18 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

MEMBRANE PROTEINSMEMGOLD THE MOST ADVANCED MEMBRANE PROTEINCRYSTALLIZATION SCREEN TO DATE.MEMBRANE PROTEINSMEMGOLD2 THE LATEST INNOVATION FOR CRYSTALLIZATION OFMEMBRANE PROTEINS. THIS SCREEN TARGETS ALLALPHA HELICAL TYPES OF PROKARYOTIC ANDEUKARYOTIC MEMBRANE PROTEINS.Crystal Growth ScreensIn 2008 Molecular Dimensions released MemGold; a rationalizedsparse matrix type membrane protein crystallization screen.MemGold was based on the crystallization conditions for 121 alphahelical Membrane Proteins deposited in the PDB.Since MemGold, the number of structures has more than doubled.In response to this, MemGold2 has been developed. Memgold2includes a further 96 crystallization conditions from unique alphahelical Membrane Protein structures including channel andtransporter structures, GPCRs and ATPases. It is suitable for bothProkaryotic and Eukaryotic alpha helical membrane proteins.Features of MemGold:n Data mined from 300 crystallization conditions for a-helicaltransmembrane proteins.n Covers conditions for approximately 130 different membrane proteinstructures in the PDB.n Addresses the diversity of membrane proteins studied.Developed by members of the So Iwata Lab, Imperial College, London.ORDER INFORMATIONDescription Pack CodeMemgold 96 x 10 mL MD1-39Memgold cacodylate-free 96 x 10 mL MD1-39-CFMemGold HT-96 96 x 1 mL MD1-41MemGold HT-96 cacodylate-free 96 x 1 mL MD1-41-CFMemGold HT-96 Green Screen 96 x 1 mL MD1-54MemGold Green Screen (10 mL) 96 x 10 mL MD1-57MemGold Combo Value Pack2 x 96 x 10 mL MD1-74MemGold Combo Value Pack HT-96 2 x 96 x 1 mL MD1-74-HTMemGold Combo Value Packcacodylate-free2 x 96 x 10 mL MD1-74-CFMemGold Screen Single Reagent 100 mL MDSR-39-MemGold HT-96 Single Reagent 100 mL MDSR-41-MemGold HT-96 Green Screen 100 mL MDSR-54-Single ReagentMemGold Green Screen Single Reagent 100 mL MDSR-57-This product is manufactured under an exclusive licence from ImperialCollege of Science, Technology & Medicine, London, UK.References1. Simon Newstead, Sébastien Ferrandon, and So IwataRationalizing α-helical membrane protein crystallization Protein Sci 2008 17: 466-472.Features of the PGA Screen:n A brand new set of 96 of the most recent alpha helical membraneprotein crystallization conditions.n Particularly suited for Prokaryotic and Eukaryotic alpha helicalmembrane proteins.n Works well in conjunction with MemGold, MemStart & MemSys& MemPlus.n Screening over a wider range of pH’s (4 - 10).n Addition of small MW PEGs.n Can be used in conjunction with Lipidic Sponge Phase and/or LipidicCubic Phases.ORDER INFORMATIONDescription Pack CodeMemGold2 (10 mL kit) 96 x 10 mL MD1-63MemGold Combo Value Pack2 x 96 x 10 mL MD1-74MemGold Combo Value Pack HT-96 2 x 96 x 1 mL MD1-74-HTMemGold2 Cacodylate-free 96 x 10 mL MD1-63-CFMemGold2 HT-96 96 x 1 mL MD1-64MemGold2 HT-96 Cacodylate-free 96 x 1 mL MD1-64-CFMemGold Combo Value Packcacodylate-free2 x 96 x 10 mL MD1-74-CFMemGold2 Single Reagent 100 mL MDSR-63-MemGold2 HT-96 Single Reagent 100 mL MDSR-64-This product was developed by Dr. Simon Newstead, University ofOxford, UK and is manufactured under an exclusive licence fromIsis Innovation Ltd.References1. Newstead, S., Ferrandon, S., and Iwata, S. ‘Rationalizing alpha-helical membrane protein crystallization‘Volume 17, Issue 3, pages 466–472, March 2008 - Protein Science, 2008 - Wiley Online Library.2. Parker, J. and Newstead, S. ‘Current trends in alpha helical membrane protein crystallization: an up-date’,Protein Science, 2012, 21 (9): 1358-1365.North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com19

Crystal Growth ScreensThe MemGold family of alphahelical membrane proteincrystallization reagents:…the science behind the screens.Dr Simon NewsteadIn recent years significant progress has been madein determining the atomic structure of alpha helicalmembrane proteins using X-ray crystallography.However the identification of conditions that givecrystals that are suitable for structure determinationremains one of the major bottlenecks.The most successful method for growing crystals of membraneproteins remains vapour diffusion from detergent solubilisedproteins. We reasoned that important advances in membraneprotein crystal screen design could be achieved by systematicallymining the information present in the Protein Data Bank and associatedresearch literature.In 2008 we undertook an analysis of the crystallization conditions for 121alpha helical membrane proteins to design a rationalized sparse matrixcrystallization screen, MemGold (Newstead et al, 2008). Our analysis hasrevealed a striking success for small MW PEGs in the crystallization ofchannels and transporters, with larger MW PEGs being more successful forrespiratory complexes and other membrane proteins that typicallycontain larger hydrophilic domains (Figure 1).3530Organic MoleculesSaltsLarge MW PEGs (3000 - 10, 000 Da)Medium MW PEGs (1000 - 2000 Da)Small MW PEGs (200 - 600 Da)No. of successful crystallisations2520151050BacterialRhodopsinsGPCRsChannelsTransportersPhotosynthetic &Light HarvestingComplexesATPasesRespiratoryComplexesOthersFigure 1. Success of small MW PEGs for crystallising alpha helicalmembrane proteins. (Reproduced with permission from Newsteadet al., Protein Science 2008).Dr Simon NewsteadInterestingly, when compared with similar analyses of successfulcrystallization data for bacterial non-membrane protein targets (Page &Stevens, 2004) from the Joint Centre for Structural Genomics in SanDiego, CA and the University of Toronto, Canada, it was clear that a majordifference is the success of small MW PEGs in promoting thecrystallization of membrane proteins. By comparison, organic moleculessuch as MPD are much less successful.20 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

50454035302520151050Small MW PEGs (200- 600 Da)Medium MW PEGs(1000 - 2000 Da)Large MW PEGs(3000 - 10, 000 Da)Figure 2. Successful crystallization precipitants reported in our2008 study for alpha helical membrane proteins compared withsimilar results reported for bacterial non-membrane proteinsfrom the Joint Centre for Structural Genomics and University ofToronto. The reported conditions in the JCSG-67 and TorontoCore-24 were found to be the smallest subset of conditions thatwould have crystallised the maximum number of bacterial nonmembraneprotein targets in these structural genomics programsin 2004.SaltsCore 67 JCSG (84 %)MemGoldCore 24 Toronto (94 %)Organic Moleculesof the crystallization conditions differed substantially, suggestingsuccessful crystallization screens should be more varied in theseparameters rather than precipitant type. Based on these data a newsparse matrix style screen, MemGold 2 was developed. Our analysis ofthe available crystallization data suggests MemGold and MemGold 2provide a comprehensive set of initial screening conditions for alphahelical membrane protein crystallization.MemAdvantage: Additive screening for membrane proteinsWe know from experience with non-membrane proteins that an initialcrystal condition will often require optimization through the addition ofsmall molecules, salts and specific ligands. To facilitate this task andagain using information gathered from the present literature onmembrane protein structures, we developed a compact, 96-reagentadditive screen, MemAdvantage. Figure 3 shows the range of differentsmall molecule and salt additives that have been reported to improveinitial crystallization conditions for alpha helical membrane proteins.Multivalent salts and polyalcohols appear prominently, accounting for10 and 15 % of all reported structures respectively. A substantialincrease in the number of secondary detergents and non-volatileorganic molecules are also now being recorded since we first startedanalyzing the data in 2008, accounting for 19 and 12 % respectively.Crystal Growth ScreensThe rate of new membrane protein structures being determined is steadilyincreasing, in line with previous predictions (White, 2004), providing anever expanding source of new crystallization information. We arecontinuing to mine this information to address the conundrum faced bymany structural biologists in this area, which is “how many conditions areenough to adequately cover ‘crystallization space’ for a given membraneprotein sample”? This question is especially pertinent for eukaryotic MPs,when amounts are likely to be far scarcer compared to their prokaryoticcounterparts. In 2012 our group undertook another detailed study of thecrystallization conditions (Parker & Newstead, 2012). Many of themembrane proteins analyzed in this study crystallized in similar precipitantconditions to our 2008 study, with approximately two thirds beingcrystallized in a low MW PEG. However, the pH, salt and buffer componentsThe rationale behind the MemGold family of membrane proteincrystallization products is to maximize the current information in thepublic databases to improve our success in crystallization and structuredetermination. Our goal in developing these screens is to provide usefulproducts to assist the structural biology community in tackling thechallenge of membrane protein crystallization, optimization andultimately structure determination.ReferencesNewstead S, Ferrandon S & Iwata S (2008) Rationalizing alpha-helical membrane proteincrystallization. Protein Sci 17: 466–472Page R & Stevens R (2004) Crystallization data mining in structural genomics: usingpositive and negative results to optimize protein crystallization screens. Methods 34: 373389Parker JL & Newstead S (2012) Current trends in α-helical membrane proteincrystallization: An update. Protein Sci 21: 1358–1365White S (2004) The progress of membrane protein structure determination. Protein Sci13: 1948–1949Figure 3. Successful additives used foralpha helical membrane proteincrystallization. The range of additivesused for successful crystallizations areshown for each membrane proteinfamily. The additives have been groupedinto more general classes for clarity andcolored according to the pie chart showninset, illustrating the composition of thecurrent database. (Reproduced withpermission from Paker and Newstead.,Protein Science 2012).North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com21

Crystal Growth ScreensMEMBRANE PROTEINSLIPIDIC-SPONGEPHASE SCREENTHIS PIONEERING SCREEN FOR MEMBRANE PROTEINCRYSTALLIZATION CONTAINS 48 CONDITIONS BASED ONPROVEN CRYSTALLIZATIONS OF TARGET MEMBRANEPROTEINS IN AN ACTIVE RESEARCH LABORATORY. 1MEMBRANE PROTEINSPGA SCREENA REVOLUTIONARY NEW SYSTEMATIC SCREEN BASED ONTHE POLY-γ-GLUTAMIC ACID (PGA) POLYMER. PGA IS THEFIRST OF THE NEW BREED OF CHEMICAL POLYMERS THATEXPLOIT POLY-AMINO ACIDS TO UNLOCK NEW AREAS OFCRYSTALLIZATION SPACE. THE PGA SCREEN IS SUITABLEFOR BOTH GLOBULAR AND MEMBRANE PROTEINCRYSTALLIZATION.The sponge phase is an expanded mesophase where the aqueouschannels are larger than in the cubic phase, thus it is able toaccommodate membrane proteins with large extra- and/or intracellulardomains. It is obtained by mixing Monoolein (MO) with abuffer solution, and a crystallization agent, such as PEG or JeffamineM600. Salt is also added to the buffer and the pH varied, allowing abroad variety of distinct sponge phases to be formed.Features of the Lipidic-Sponge Phase Screen:n Guaranteed lipidic-sponge phases, premixed and ready-to-use!n Closely resembles the native lipidic environment of membraneproteins.n Suitable for use in hanging or sitting-drop vapour diffusionexperiments, and in lamina (p.73).n Easy to handle liquids.n Easy retrieval of protein crystals.n No lipase or cryoprotectantsneeded.n Can be used with additive andcrystallization screens.n Easy to spot colourless crystals.n Readily compatible withhigh-throughput crystallizationapproaches.The high nucleation-precipitation potential of PGAs enables theiruse at very low concentrations and in combination with classicalprecipitants, thus scaling down the amount of precipitantnecessary for crystal appearance and growth. This feature of PGAsmakes them especially useful in applications for labile, easilyprecipitating proteins. PGA Screen is suitable for both globular andmembrane protein crystallization!Features of the PGA Screen:n Novel precipitant.n Totally New Crystallization Space.n Easy mixing properties with other PEGs.n Applicable to both globular and membrane protein crystallization.n Especially useful for labile, easily precipitating proteins.n Non-toxic and non-denaturing.n Compatible with liquid – handling robots.ORDER INFORMATIONDescription Pack CodeThe Lipidic-Sponge Phase Screen 48 x 40 µL MD1-48References1. Annemarie B. Wöhri, Linda C. Johansson, Pia Wadsten-Hindrichsen, Weixiao Y. Wahlgren, Gerhard Fischer,Rob Horsefield, Gergely Katona, Maria Nyblom, Fredrik Öberg, Gillian Young, Richard J. Cogdell, Niall J.Fraser, Sven Engström, Richard Neutze. “A lipidic-sponge phase screen for membrane proteincrystallisation” Structure, Vol 16, 1003-1009 (2008).ORDER INFORMATIONDescription Pack CodeThe Lipidic-Sponge PhaseReservoir Solution 100 mL MD2-100-85The Lipidic-Sponge PhaseReservoir Solution 250 mL MD2-250-85Monoolein 500 mg MD2-67Monoolein 1G MD2-68Lipidic-Cubic Phase Mixing Adapter 40 µL MD6-17The Lipidic-Sponge PhaseSingle Reagent 40 µL MDSR-48-PGA Screen premixed with buffers 96 x 10 mL MD1-50PGA Screen premixed with bufferscacodylate-free 96 x 10 mL MD1-50-CFPGA Screen HT-96 96 x 1 mL MD1-51PGA Screen HT-96 cacodylate-free 96 x 1 mL MD1-51-CFPGA screen PREMIXED Single Reagent 100 mL MDSR-50-PGA screen PREMIXED HT-96Single Reagent 100 mL MDSR-51-This product is manufactured under an exclusive license from YorkStructural Biology Laboratory, University of York, UK.References1. TC Hu, J Korczynska, DK Smith, AM Brzozowski - Acta Crystallographica Section D: BiologicalCrystallography, 2008. D64, 957-963.22 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

MEMBRANE PROTEINSMEMMAGIC ®BICELLE SCREEN KITTHE MEMMAGIC BICELLE SCREEN KIT IS BASED ON THEUSE OF BICELLES AS AN ALTERNATIVE METHOD FOR THECRYSTALLIZATION OF MEMBRANE PROTEINS IN A LIPIDICENVIRONMENT.MEMBRANE PROTEINSTHE CALIXAR ADDITIVE KITTHIS KIT IS DESIGNED TO TEST THE EFFECT OF 24 NEWCOMPOUNDS ON PROTEIN CRYSTAL GROWTH.Crystal Growth ScreensMembrane proteins can be readily reconstituted into bicelles andare maintained in a native-like bilayer environment, which can bemanipulated with almost the same ease as for detergent solubilizedmembrane proteins, making it compatible with standard highthroughputscreening methods.Features of the MemMagic ® Bicelle Screen Kit:n Handles like detergentn Behaves like lipidn Robotically compatibleApplications:n Crystallographyn NMRn Assay DevelopmentMemMagic ® Bicelle Screen kit provides four bicelle solutions of 40%,35%, 30%, 25% DMPC:CHAPSO (2.8:1).New reagents have been designed to structure the membranedomains through hydrophobic interactions and a network of saltbridges with the basic residues found at the cytosol-membraneinterface of membrane proteins. These reagents behave assurfactants forming micelles of 5–24 nm, with the critical micellarconcentration (CMC), (being as expected sensitive to pH), rangingfrom 0.05 to 1.5 mM. Such molecules were successfully used topromote crystallization of membrane proteins. The figure shows anexample of 12 transmembrane protein (ABC transporter) that wasconcentrated to 10mg/mL, crystallized using the sitting drop vapourdiffusion method in 0.2M KSCN, 20% PEG 3340 in the presence ofthese new compounds. This example illustrates that these additivescan promote membrane protein crystallization and help to obtaindiffracting protein crystals.Features of The Calixar Additive Kit:n Promotes crystallization of functional membrane proteinsn Generates a saltbridge network around the protein and hydrophobiccontactsThis product is manufactured and distributed under an exclusive licenseusing CALIXAR patented technology and is supplied for research useonly. This product may not be used for the provision of commercialservices for which a separate license must be obtained.Bicelles are disk-like micelles formed by the mixture of a phosphatidylcholine lipid such as Dimyristoylphosphatidylcholine(DMPC) and a detergent such as 3-[(3-cholamidopropyl) dimethylammonio]-2-hydroxy-1-propanesulfonate (CHAPSO). The bicelle discs can be described as patches of lipid bilayers with detergentmolecules lining the apolar edges of each bilayer.ORDER INFORMATIONDescription Pack CodeMemMagic ® Bicelle Screen kit 4 x 100 µL MD1-81MemMagic ® Bicelle Screen kit 4 x 250 µL MD1-82MemMagic ® and MemX ® are registered trademarks ofMemxBiosciences LLC.ReferencesFaham, S., Ujwal, R., Abramson, J. and Bowie, J. U. (2009) “Practical Aspects of Membrane ProteinsCrystallization in Bicelles” Current Topics in Membranes, Volume 63, Chapter 5, 111-127.Faham, S., Boulting, G. L., Massey, E. A., Yohannan, S., Yang, D., & Bowie, J. U. (2005). “Crystallization ofbacteriorhodopsin from bicelle formulations at room temperature”. Protein Science, 14, 836–840.Faham, S., & Bowie, J. U. (2002). “Bicelle crystallization: A new method for crystallizing membrane proteinsyields a monomeric bacteriorhodopsin structure”. Journal of Molecular Biology, 316, 1–6.Figure: CALIXAR compounds promote crystallization of a 12 transmembrane target protein (ABC Transporter)only in presence of these compounds at different concentration.ORDER INFORMATIONDescription Pack CodeThe Calixar Additive Kit 24 x 50 µL MD1-80CALIXAR Single Reagent 50 µL MDSR-80-CALIXAR single reagents are manufactured by Calixar SAS andexclusively licensed to Molecular Dimensions for the manufacture anddistribution of this additive kit for final end users only. The use of this kitfor commercial service purposes requires a separate licence fromCalixar SAS.ReferencesMatar Merheb M et al., PLoSONE 2011North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com23

Crystal Growth ScreensMEMBRANE PROTEINSMEMADVANTAGE AN ADDITIVE SCREEN DEVELOPED EXCLUSIVELY FOR MEMBRANE PROTEINS. THISSCREEN TARGETS ALL ALPHA HELICAL TYPES OF PROKARYOTIC AND EUKARYOTICMEMBRANE PROTEINS. MEMADVANTAGE WAS DEVELOPED FROM THEIDENTIFICATION OF SUCCESSFUL ADDITIVES (USING DATA MINING) CURRENTLYUSED IN THE CRYSTALLIZATION OF MEMBRANE PROTEINS. IT CONTAINS A NOVELSET OF CHEMICALS PRESENTED AS A 96-FORMAT SCREEN FOR IMPLEMENTATIONIN ROBOTIC SCREENING PIPELINES.The kit is designed to help test the effect of 96 different compoundson membrane protein crystal growth. Detergent selection is a criticalparameter for growing well-ordered, well diffracting crystals and withso many choices of detergents/ligands to choose it can be both timeconsuming and expensive to investigate all possibilities.MemAdvantage takes the most successful ligands, detergents,multivalent salts, polyalcohols, non-volatile organics, organics,amphiliphiles and puts them all together in one easy-to-use additivescreen. Additives may affect hydration and intermolecularinteractions between protein molecules or between protein moleculeand solvent and even ligands. This kit is a screen and results mayneed to be interpreted with a view to designing further additiveexperiments using different compounds of the same type as the kitreagent that gave a promising result.ORDER INFORMATIONDescription Pack CodeMemAdvantage HT96 x 0.25 mL MD1-70For MemAdvantage reagents see page 49.Features of MemAdvantage:n A rational and intelligently designed additive screen targetedspecifically for membrane proteins.n Allows easy screening of 96 different additives (12 different classes ofthe following: polyalcohols, detergents, multivalent salts, non-volatileorganics etc.) found to be the most successful in membrane proteincrystallization.n Particularly suited for Prokaryotic and Eukaryotic alpha helicalmembrane proteins.n For initial screening or optimization screening.n Ready-to-use deep-well block.ReferencesParker, J. and Newstead, S. ‘Current trends in alpha helical membrane protein crystallization: an update’,Protein Science, 2012, 21 (9):1358-1365.This product is manufactured under an exclusive licence from IsisInnovation Ltd, and was developed by Dr. Simon Newstead, Universityof Oxford, UK.Additives found inMemAdvantage and theirsuccessfulness incrystallization ofmembrane proteins.24 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

NUCLEIC ACIDNUCLEIC ACIDHELIX – THE LATEST INNUCLEIC ACID SCREENINGHELIX - PROVIDES A VARIETY OF APPROACHES TOENHANCE SUCCESSFUL CRYSTALLISATIONS FOR ADIVERSE RANGE OF NUCLEIC ACID TOPOLOGIES ANDMOLECULAR WEIGHTS.HELIX is a 96 condition nucleic acid focused kit designed tomaximize the diversity of conditions, and is modular in its design toallow for focused experiments. Conditions include: a 24-well subsetcovering cryo-cooling; a 24-well subset covering four-strandedDNA/RNA quadruplexes; a 12-well subset focused on low pH fori-motifs, and cytosine protonation; a 12-well subset focused onlarge molecular weight nucleic acids; a 24-well subset coveringdivalent metal ions and additives designed to be combined invarious ways to rapidly generate customised orthogonal screens.Crystal Growth ScreensFeatures of HELIX:n Nucleic acid fragments, of all molecular weightsn Double stranded DNA and RNA, pseudo knots, G-quadruplexes,i-motifs, triplex, ribozymesn Optimized for MAD, SAD, SIRAS data collectionsn Cryo-cooling optimizationn Screening of additivesn Adaptable for High Throughput ScreeningORDER INFORMATIONDescription Pack CodeHELIX 96 x 10 mL MD1-68HELIX HT-96 96 x 1 mL MD1-69HELIX single reagent 100 mL MDSR-68-HELIX HT-96 single reagent 100 mL MDSR-69-Developed by Dr. Gary Parkinson from School of Pharmacy, UniversityCollege, London, UK and manufactured under an exclusive license fromthe University of London.North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com25

OPTIMIZATIONTO OPTIMIZE INITIAL HITS FROM SCREENING, MOLECULAR DIMENSIONSPROVIDES A COMPLETE RANGE OF BUFFERS, CRYSTALLIZATION STOCKSOLUTIONS AND SINGLE SCREEN REAGENTS.30 Custom screens31 Naomi’s nucleant31 MicroSeed Beads35 Fluorescent labelling kits36 Additive screen36 Single reagents37 PEG precipitants38 Alternative precipitiants39 Jeffamines ®39 Volatiles – organics40 Non-volatiles – organics40 Polyamines41 Salts43 Other reagents43 Cryoprotectants44 Buffer stocks45 Special buffers45 Morpheus ® mixes46 Morpheus ® individualstock reagents48 Midas reagents49 MemAdvantage reagents51 The Really Useful Buffer kit51 Gelled Surface Kit51 Microbatch Oils26 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

SEEDINGLABEXERCISESDr Terese Bergfors1. What is seeding?• The use of an existing nucleus (usually a small crystal or crystalfragment), introduced into a new drop, where it acts as a growth site.2. Why seed?• To separate nucleation from growth and bypass the need forspontaneous nucleation. It is easier to add onto an already existingnucleus than create one de novo. See Fig. 1. Seeding and the phasediagram.• Improve the size of the crystals or control the number of crystals• Get more consistent results when crystals don’t always appear inknown conditions• Speed up results if spontaneous nucleation is slow• To obtain a wider range of crystal forms (polymorphs) by seedinginto totally different precipitantsIn this article we will present twolaboratory exercises:1. Instantaneous streak seeding varying theprotein concentration.2. Comparison of jab and streak seeding withovernight equilibration.3. What are the types of seeding?• Macroseeding is the transfer of a single, pre-grown, washed crystal.• Microseeding is the transfer of microscopic crystals, crushed up intofragments.• Streak seeding is a form of microseeding that transfers themicroseeds by a stroking motion with a whisker or hair of somesort. See Fig. 2. Streak seeding. Can be used on its own or incombination with microseeds in a dilution series.• Jab seeding is another variationon microseeding where the newdrop is “innoculated” with a single jab of the seed transfer tool.Reference:For a review article on seeding, see: Bergfors, T. “Seeds to Crystals”J. Structural Biol. 2003, vol. 142, 66-76North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com27

WaxProbe[Macromolecule][Crystallizing agent]UndersaturationSaturationNo seed crystal addedMetastable zoneLabile zoneSeed crystal addedPrecipitation zoneFigure 1: Seeding and the phase diagram.From Luft and DeTitta, Acta Cryst. (1999) D55, 988-993.Figure 2: Streak seeding from Enrico Stura, Published in “Crystallization ofNucleic Acids and Proteins” Edited by Ducruix and Giege, 1992, OxfordUniversity PressEXPERIMENT 1:Instant streak seedingReference: This exercise has been adapted from Enrico Stura’s chapter"Seeding" in Protein Crystallization: Strategies, Techniques, and Tipsedited by T. Bergfors 1999 International University Line, La Jolla Ca.Purpose of this experiment:1. To learn how to generate new seeds by the easy, fast, and simplemethod of streak seeding.2. To observe the effects of decreasing protein concentration on thenucleation rate.Materials needed:• A crystal wand – you can make your own from a cat whisker, humanhair or horse tail hair, super glue or wax to affix the whisker to ayellow pipette tip, and use a razor blade to cut the whisker.• lysozyme 100 mg/mL stock solution.• A dilution series of lysozyme at 80, 60, 40 and 20 mg/mL.• 30% polyethylene glycol (PEG) 5000 or 6000 in 1 M NaCl, 50 mMNa acetate, pH 4.7 = the precipitant.• A surface for making the drops, e.g., a Petri dish lid, cover slip, etc.Method:1. On the Petri dish lid, pipette 10 µl of lysozyme (100 mg/mL). Now addan equal volume (i.e., 10 µl) of the precipitant (= 30% PEG 6000 in1M NaCl, 50 mM Na acetate, pH 4.7).2. Watch the drop under the microscope. The first thing you should seeis a phase separation. After that, the crystals should nucleate in5-15 minutes. The fresher the lysozyme the longer time it will taketo nucleate, so the time will vary. It can take some practice torecognize the nucleation in its initial stages. Note: If the proteinprecipitates immediately, the protein concentration is too high.Redo with a slightly lower protein concentration; try 80 mg/mL.3. Once the parent crystals have formed in the drop with 100 mg/mL, youcan set up the 10 µl drops of lysozyme at 80, 60, 40, and 20 mg/mLon the inside of a Petri dish lid.100 mg/ml4. Add 10 μl of the precipitant to each of these 4 new drops and stir.Important! These drops also need precipitant, not just protein, or theexperiment does not work. Students often forget this step.5. Now you can transfer the nuclei (from the parent drop) by streakseeding. Dip the seeding wand once into the drop of parent crystals topick up the seeds, then streak the wand across the four new drops. Youdo not need to re-dip the wand into the parent drop for each new drop.What to look for or think about:• See if you can get the crystals to grow along a streak line. Because ofthe high protein concentrations you will also get spontaneousnucleation but many crystals will form preferentially along the streakline. The crystals will be extremely small because they grow so rapidly.In this experiment the protocol has been optimized for speed, ratherthan crystal size.• You don't need to save these drops, but you can use them in the nextexperiment. In the current experiment we demonstrated the methodand the principles for streak seeding. In the next experiment,a more refined version will be presented.28Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

EXPERIMENT 2:Comparing the effects of addingdifferent amounts of seedsPurpose of this experiment:1. To illustrate the effect of adding different amounts of seeds.2. To illustrate a very fast and easy way of creating a dilution series ofseeds.In contrast to experiment 1 left, this experiment will be allowed toequilibrate overnight to permit slower growth of the crystals after seedsare added.Three rows of identical drops will be prepared. Seeds will beadded by either jabbing the drops or streak seeding them. These dropswill be compared to a control where no seeds have been added. Anentire row of drops will be seeded sequentially to create a dilution of theseeds. Thus, the first drop will have the most seeds and the last drop inthat row will have the least.Materials required:• Two 20 mL Petri dishes (the large size)• 5 mL of 6% NaCl in 50-100 mM sodium acetate buffer, pH 4.7• 5 mL of 12% NaCl in 50-100 mM sodium acetate buffer, pH 4.7• 100 microliters of 20 mg/mL lysozyme, dissolved in water• A seeding wand (you can use the same one from the previousexperiment.)• Two Eppendorf tubesMethod:1. In an eppendorf tube, mix 50 microliters of lysozyme (20 mg/mL)with 50 microliters of 6% NaCl in 50-100 mM sodium acetate buffer,pH 4.7 = MIXTURE A.2. In another eppendorf tube (mark them, so you don’t mix them up!),mix 50 microliters of lysozyme (20 mg/mL) with 50 microliters of 12%NaCl in 50-100 mM sodium acetate buffer, pH 4.7 = MIXTURE B.3. In one Petri dish, pour approximately 5 mL of the buffered 6% NaClsolution in the bottom = PLATE A.4. Repeat for the other Petri dish, using buffered 12% NaCl instead =PLATE B.5. Using MIXTURE A, pipette 3 rows of 6 drops each onto the lid ofPLATE A. Each drop will contain 5 microliters. See picture:6. Repeat for MIXTURE B and PLATE B.7. Row 1 will be the control.8. Row 2 will be jab seeded. Using the seed transfer tool, dip or stir itinto the parent drop in the previous experiment (experiment 1) topick up seeds.9. Now jab all the drops in row 2 with the seed transfer tool. To do so,barely touch the outermost edge of the drop with the tool = a quickjab. Rinse the tool in the reservoir solution between each jab tocreate a dilution series of the seeds.10. Row 3 will be streak seeded. Rinse the seed transfer tool thoroughlyand wipe it off with a tissue. Dip or stir it into the parent drop in theprevious experiment (experiment 1) to pick up a new batch of seeds.11. Now streak seed the drops in row 3 with the seed transfer tool. Rinsethe tool in the reservoir solution between each streak to create adilution series of the seeds.12. Invert the lids of the Petri dishes over the correct, respectivereservoirs and seal the rims with parafilm.13. Wait 24 hours to observe the results.Expected results:1. After 24 hours, the control row in PLATE A will probably be still clear(but crystals will grow there after 48 hours, so it is best to examinethe plates before that). In PLATE B, you probably will see sea urchintypecrystals by 24 hours.2. The rows with jab seeding and streak seeding will give differenteffects. Jab seeding works best if the drop is very large, becausethere is a dilution effect across the surface of the drop. The dropshere are only 5 microliters. Once back home, try the effect on largesitting drops (40 microliters) to make it more pronounced.3. Jab and streak seeding several drops in a row (serial seeding) is a fastand easy way of making a dilution series of seeds in the new drops.The drops seeded last (by either method) should show the leastnumber of crystals compared to the first drops in the row. See anexample in Figure 3.control rowjab seedstreak seedFigure 3. Comparison of drops that have been serially seeded with a control.These results were photographed 48 hours after setup. Row 1 is a control andhas not been seeded. The lysozyme crystallizes in the form of sea urchins orspherulites. Row 2 has been jab seeded. The needle inoculated with seeds wasjabbed into the center of the drops. Row 3 has been streak seeded. Notice thatcrystals tend to form along the seed line, but there is much spontaneousnucleation even outside the streak line in the first drops. However, the final dropin the row (drop 5 in the series) has nice, large crystals. This effect was achievedbecause the number of seeds added to the drop was diluted by passing thestreak wand through the first drops in the row. (Photos by Terese Bergfors).North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com29

Optimization48 & 96 REAGENTCUSTOM SCREENOver the years Molecular Dimensions havegained extensive experience of producingcustom crystallization screens for its manycustomers. (Some of these screens have sincegone on to become some of our best-sellingcrystallization screens, (e.g. Morpheus ® , ClearStrategy & MemGold).We can now provide custom crystallizationscreens & solutions designed by you. Simplyprovide the specifications; molarity, pH etc.and we’ll do the rest; from small-scale (mL) tolarge-scale (L).All solutions are manufactured to our exactingstandards using ultra-pure 18 MegaOhm water.All custom reagents & screens are treated asstrictly confidential and are provided withfull traceability.CONTACT MOLECULAR DIMENSIONSFOR YOUR TAILORED CUSTOMSCREEN QUOTATION“I have been ordering customscreens from Molecular Dimensionssince I started my crystallographyproject 3 years ago. Very quickand convenient !!!”MRC Prion Unit, London, UK.ORDER INFORMATIONDescriptionCode48 reagent Custom screen Set-up+ 1 kit (10 mL or HT96) MD1-CUSTOM4848 reagent custom screen10 mL kit ( 1 box) MD1-CUSTOM48-10ML48 reagent custom screen HT-96 kit MD1-CUSTOM48-HT9696 reagent Custom screen Set-up+ 1 kit (10 mL or HT96) MD1-CUSTOM9696 condition custom screen10 mL kit (2 box kit) MD1-CUSTOM96-10ML96 condition custom screen HT-96 kit MD1-CUSTOM96-HT9630Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

NUCLEANTNAOMI’S NUCLEANTNAOMI’S NUCLEANT HAS FACILITATED THECRYSTALLIZATION OF THE HIGHEST NUMBER OFPROTEINS REPORTED FOR ANY SINGLE NUCLEANT.MANY OF THESE PROTEINS HAVE PROVEN DIFFICULTTO CRYSTALLIZE AND SOME OF THESE, INCLUDINGMEMBRANE PROTEINS, HAVE ONLY BEEN CRYSTALLIZEDIN THE PRESENCE OF NAOMI’S NUCLEANT.In addition to test proteins those proteins that can be named are:multi drug resistance protein (a membrane protein), modifiedcyclodextrine, oxyntomodulin, myosin binding protein C, lobstershell α-crustacyanin, c-phycocyanin, α-actinin actin bindingprotein. Several other proteins have also been crystallized butcannot be named at this time. Often the crystals obtained were ofincreased diffracting quality compared to those resulting fromstandard techniques. For example myosin binding protein Cdiffracted to 1.6Å compared to 3Å.Features of Naomi’s Nucleant:n Simply add a single grain to a crystallization drop.n Easy to place with fine tweezers.n Use in screening or optimization to nucleate supersaturatedconditions.n Use in optimization where excessive nucleation occurs (i.e. lots of tinycrystals). Back off the precipitant concentration to the metastablezone and then use a grain to nucleate.n Negates twinning.n Protein crystals are easily detached from the nucleant using amicroprobe or a cryo loop.SEEDINGMICROSEED BEADS FOR THE PRODUCTION OF MICROSEEDS FOROPTIMIZATION EXPERIMENTS.MicroSeed Beads are used to generate microseeds (submicroscopiccrystals that have been ‘crushed-up’ into fragments).Crystals are crushed using PTFE balls and suspended into serialdilutions of mother liquor.Features of MicroSeed Beads:n Improve the size of crystals or control the number of crystals.n Get more consistent results.n Speed up results if spontaneous nucleation is slow.n Avoid cross-contamination unlike glass beads.n Obtain a wider range of crystal forms (polymorphs) by seeding intototally different precipitants.ORDER INFORMATIONFind out more aboutMicroSeed Beads hereDescription Pack CodeMicroSeed Beads 24 x PTFE MicroSeed Beads MD2-14In individual 1.5 mLmicrofuge tubesOptimizationPHASINGI3C PHASING KITORDER INFORMATIONCrystals (arrowed) of β-lactamase grownon a grain of Naomi’s Nucleant by RosalidaLeone at Imperial College, London.Description Pack CodeNaomi’s Nucleant 1 vial MD2-07(approx 3mg)(approx. 300 grains)Microspatula 1 MD9-08Fine tweezers 1 MD9-25Microprobe sampler kit 1 kit MD9-01Developed and patented at Imperial College London, the most effectivenucleant of any material tested.References1. Chayen, N.E., Saridakis, E. and Sear R. Experiment and theory for heterogeneous nucleationof protein crystals in a porous medium. PNAS (2006) 103, 597-601.2. Saridakis, E. and Chayen N.E., Towards a ‘universal’ nucleant for protein crystallization.Trends in Biotechnology (2009) 27, 99-1063. Eisenstein, M. The shape of things. Nature Methods (2007) 4, 95-101.FOR HEAVY ATOM DERIVITIZATION OF BIOLOGICALMACROMOLECULES FOR SUBSEQUENT SINGLEWAVELENGTH ANOMALOUS DISPERSION (SAD) ORSINGLE ISOMORPHOUS REPLACEMENT WITHANOMALOUS SCATTERING (SIRAS)Features of I3C Phasing Kit:n Use in heavy atom soaking experiments.n Use in co-crystallization.n Pre-weighed aliquots.ORDER INFORMATIONDescription Pack CodeI3C Phasing Kit 12 aliquots of I3C (280 mg MD2-11each) + 12 aliquots of 2.0 MLiOH (650 µL each)North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com31

TRACE FLUORESCENTLABELLING USINGVISIBLEFLUORESCENCE FORCRYSTALLIZATIONSCREENINGDr Marc PuseyINTRODUCTION:Obtaining sufficiently well diffracting crystals is the bottleneckprocess in macromolecule structure determination. Findingthese conditions depends upon determining successfuloutcomes in screening trials. Current trends are to utilize eversmaller crystallization trial volumes, to enable setting up more trials.This results in large numbers of experiments to be set up, maintained, andperiodically monitored, leading in turn to the increased use of robotics forall aspects of the procedure. Robotic analysis of white light images is notsimple, and is liable to be confounded by, e.g., masses of crystals orcrystals buried in precipitate. The latter outcome can also presentproblems to a manual visual analysis of crystallization trials, a processthat can result in missing many leads and also a mind-numbingly tedioustask for even small scale screening trials and the results therebynegatively affected.These considerations led to development of the trace fluorescent labelingapproach for crystallization screening (Forsythe et al., 2006). The basisof this method is that

As the emitted intensity is a function of the fluorescent probeconcentration, and the crystalline state is the most densely packedfor the (labeled) protein, then crystals are the brightest objects in theobserved field. Automatically acquired image data, as thumbnails, can berapidly scanned and crystals or likely lead conditions quickly picked out.This monograph describes the trace fluorescent labeling procedure,gives examples of outcomes using this procedure, and briefly describesthe potential applicability of this method for more difficult crystallizationproblems.Labelling:There are three approaches to the use of fluorescence in proteincrystallization screening. The first and most direct is to use the intrinsicfluorescence of the protein (Judge et al., 2005) (see Green Screens). Thisrequires a UV light source, UV transmissive optics, and an imaging systemsensitive in the UV, in all cases considerations that add cost and safetyconcerns to this method. Additionally, other biological compounds mayfluoresce and be a source of interference. As well, there are often safetyconcerns associated with the use of UV light. The second is to add aprobe to the solution that is non-fluorescent until bound to ahydrophobic region (Groves et al., 2007). This is also a simple and directmethod, with excitation in the near UV (375 nm) and emission in thevisible (~460 nm). In our hands we find that it is considerably less sensitivethan the third approach, the trace (

What to look for “Intensity = Structure”The premise behind the trace labeling approach is that the observedfluorescence intensity is proportional to the density of fluorescingmolecules. As the crystalline state is the most dense for any materialunder normal circumstances then crystals will fluoresce the mostintensely. Thus we get the primary maxim for using fluorescence;intensity equals structure. Optimal probe concentrations are in the ≤1 x 10 -6 M range. A quick calculation indicates that most crystallineprotein concentrations are in the millimolar concentration range, andthus at 0.1% labeling are at good concentrations for detection. Figure1 shows images of a 3D crystal and needles using trace fluorescentlabeling and white light imaging. Figure 2 shows a drop with crystalsand precipitate. Note the ease with which the crystals can beobserved as a result of their greater intensity with respect to theprecipitate.Figure 1: Fluorescent and white light images of a 3D crystal (top row) and needles (bottom row).Choice of Fluorescent ProbeAn advantage of using visible fluorescence is the wide range of coloursone has to choose from, making it easy to avoid potentially troublesomeinterfering fluorescence. Our first choice fluorescent probe isCarboxyrhodamine, which excites at ~530 nm and emits at ~550 nm,close to the maximum sensitivity for the human eye.This probe also has a very high absorptivity and quantum efficiency.Other advantages, which should be considered when selecting otherprobes, are that it is relatively insensitive to solution factors such as pH.Fluorescence can be a very sensitive reporter of the probes immediateenvironment. Given that a crystallization screen introduces a wide rangeof conditions it is preferable to use a probe that minimizes thisenvironmental sensitivity for a consistent signal over the broad range ofconditions to be expected.Observing the crystallization platesCrystallization plates can be viewed traditionally using a low poweredwhite light microscope. For fluorescent viewing one should use afluorescent microscope with a low power (≤4X-5X) objective. Theexcitation and emission filters should be appropriate for the probeemployed. Alternatively, one can use a plate imaging system designedfor use with visible fluorescence, again with the appropriate filters.*For safety reasons it is NOT recommended that one use a low powermicroscope with, e.g., direct laser excitation impinging upon the drop, asreflected laser light could damage the eye. We have also tried usingdirect excitation using light emitting diodes at the appropriatewavelength, without any filters, to no avail.The excitation and emission filters are necessary to remove overlappingexcitation light from the emission light.*Please refer to the Crystal X2 (page 126).An advantage of fluorescence is that one can also derive cryptic leadconditions from the results. Using the maxim that intensity equalsstructure, we have often found drops showing intensity that did notcorrespond to any discernible structure upon close examination ofthe drop under white light. Optimization screening around theseconditions often results in crystals, as shown in Figure 3. Theprecipitate in panel A shows bright regions in the fluorescent image,which were not resolvable as discrete structures in the white lightimage. The solution in panel A showed nothing in the white lightimage, but had bright regions under fluorescence. The “structure” inpanel C did not appear crystalline, but note that the fluorescenceimage shows a clearing zone around it, suggesting that it is growing by‘feeding’ off of the surrounding precipitate. Notice that this additionalclue is not obvious in the white light image. In all three cases, clues asto the behavior of the protein in the solution are derived from carefulanalysis of the results, and subsequently acted upon to producecrystals. The optimization in all three cases was carried out bycapillary counter diffusion, as shown in the lower images. From left toright the capillaries have internal diameters of 200, 160, and 100 µm.Use with complexesAn advantage of using visible light trace fluorescent labelling is thatone can employ multiple colors. This makes the methodology veryuseful for avoiding potentially interfering materials that may bepresent, and for identification of the crystallization of complexes. Forcomplexes one can trace label different components with a differentcolor, and then use the fluorescence to determine if all componentsare present in any crystals found upon screening. The crystallineprobe concentrations are such that the average separation betweenprobe molecules is > than the distances that would result inresonance energy transfer, obviating this as a concern in the imagingprocess. Figure 4 shows a test case, where two different solutions of asingle protein were labeled with different fluorescent probes, mixedtogether, and then crystallized. As a control, plates were alsoseparately set up with each labeled protein.References:Forsythe, E., Achari, A., and Pusey, M. L. (2006). Trace fluorescent labeling for high-throughputcrystallography. Acta Cryst. D 62:339-346.Groves, M. R., Müller, I. B., Kreplin, X., and Müller-Dieckann, J. (2007). A method for the generalidentification of protein crystals in crystallization experiments using a noncovalent fluorescent dye.Acta Cryst. D 63:526-535.Judge, R. A., Swift, K., and Gonzalez, C. (2005). An ultraviolet fluorescence-based method for identifyingand distinguishing protein crystals. Acta Cryst. D 61:60-66.Pusey, M., Forsythe, E., and Achari, A. (2008). Fluorescence approaches to growing macromoleculecrystals. Meth. Mol. Biol. 426:377-385.34 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

REAGENTSTHE TRACE FLUORESCENCELABELLING KITOptimizationTHE BRILLIANT ANSWER TO YOUR CRYSTALLIZATIONPROBLEM. A SIMPLE, FAST AND FLEXIBLE LABELLINGKIT - LABEL HAS NO PROVEN EFFECT ON THE PROTEIN,CRYSTALLIZATION AT STIPULATED LABELLING LEVELSOR DATA QUALITY.Figure 2: Fluorescent (left) and white light images of crystals buried in precipitant. While the precipitantfluoresces, the intensity from the crystals is significantly greater, rendering them easy to see.Label two components with different colour dyes - uniquely checkboth parts of a complex are in the crystal.Features of The Trace Fluorescence Labelling Kit:n Significantly higher and faster crystal identification rate.n Ideal for verifying the presence of complexes (requires dual labellingkit MD1-72).n Stunning signal-to-noise ratio provides significantly higher crystalidentification rate.n Provides clear views through precipitate.n No false positives or negatives.n

OptimizationREAGENTSADDITIVE SCREENTHIS ADDITIVE SCREEN SAMPLES THE VARIOUSCLASSES OF TYPICAL ADDITIVES USED TO OPTIMIZECRYSTALLIZATION.REAGENTSSINGLE REAGENTSSINGLE REAGENTS FROM SCREENS ARE AVAILABLE IN100 ML QUANTITIES.As such it is an inexpensive alternative to screening a large numberof compounds. Positive results achieved with one or more of theseadditives could be further explored by testing other compounds fromthe same class.All of the single conditions from Molecular Dimensions’ crystalgrowth screens are available as single reagents in 100 mL quantities.Features of the Additive Screen:n Screen classes of typical additives.n An excellent starting point to improve the diffraction quality,morphology or size of a protein crystal.Features of Single Reagents:n Made to order.n Prepared to the same quality standards as the parent screen.n Prepared using the same water and chemicals used in the screens.n Sterile filtered.ORDER INFORMATIONDescription Pack CodeAdditive Screen 24 reagents MD1-11ORDER INFORMATIONTo order please quote code as follows:Code: MDSR-Screen Code - (Box No.) – tube or well numbere.g. Reagent 5 from Structure Screen 1 (MD1-01) = MDSR-01-5;Reagent 17 from box 2 of Proplex (MD1-38) = MDSR-38-2-17;Reagent B4 from ProPlex HT-96 (MD1-42) = MDSR-42-B4.36 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

REAGENTSPEG PRECIPITANTSSTOCK SOLUTIONS OF PEGS AND PEG MME'S.AVAILABLE IN 100 ML OR 250 ML QUANTITIES.OptimizationORDER INFORMATIONDescription Concentration Code Available volumesPEG 200 100% MD2-100(250)-1 100/250 mLPEG 300 100% MD2-100(250)-2 100/250 mLPEG 400 100% MD2-100(250)-3 100/250 mLPEG 600 50% v/v MD2-100(250)-4 100/250 mLPEG 1000 50% w/v MD2-100(250)-5 100/250 mLPEG 1500 50% w/v MD2-100(250)-6 100/250 mLPEG 2000 50% w/v MD2-100(250)-7 100/250 mLPEG 3000 50% w/v MD2-100(250)-8 100/250 mLPEG 3350 50% w/v MD2-100(250)-9 100/250 mLPEG 4000 50% w/v MD2-100(250)-11 100/250 mLPEG 6000 50% w/v MD2-100(250)-12 100/250 mLPEG 8000 50% w/v MD2-100(250)-13 100/250 mLPEG 10,000 50% w/v MD2-100(250)-14 100/250 mLPEG 12,000 50% w/v MD2-100(250)-15 100/250 mLPEG 20,000 50% w/v MD2-100(250)-16 100/250 mLPEG 350 MME 100% MD2-100(250)-18 100/250 mLPEG 500 MME 100% MD2-100(250)-66 100/250 mLPEG 750 MME 50% v/v MD2-100(250)-67 100/250 mLPEG 2000 MME 50% w/v MD2-100(250)-17 100/250 mLPEG 5000 MME 50% w/v MD2-100(250)-19 100/250 mLNorth & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com37

OptimizationREAGENTSALTERNATIVE PRECIPITANTSSTOCK SOLUTIONS OF ALTERNATIVE PEG-BASED PRECIPITANTS.AVAILABLE IN 100 ML OR 250 ML QUANTITIES.ORDER INFORMATIONDescription Concentration Code Available volumesAcrylic acid/maleic acid copolymer (50:50), Na salt 35% v/v MD2-100(250)-130 100/250 mLCetyltrimethylammonium bromide (CTAB) 0.05M MD2-100(250)-28 100/250 mLGlycerol ethoxylate 50% v/v MD2-100(250)-132 100/250 mLPentaerythritol ethoxylate (3/4 EO/OH) 50% v/v MD2-100(250)-141 100/250 mLPentaerythritol propoxylate (5/4 PO/OH) 50% v/v MD2-100(250)-115 100/250 mLPentaerythritol ethoxylate (15/4 EO/OH) 50% v/v MD2-100(250)-140 100/250 mLPentaerythritol propoxylate (17/8 PO/OH) 50% v/v MD2-100(250)-142 100/250 mLPoly-glutamic acid (PGA)-LM 20% w/v MD2-100(250)-108 100/250 mLPoly-glutamic acid (PGA)-HM 20% w/v MD2-100(250)-162 100/250 mLPolyacrylate 2100, sodium salt 50% w/v MD2-100(250)-143 100/250 mLPolyacrylate 5100, sodium salt 50% w/v MD2-100(250)-144 100/250 mLPolypropylene glycol 400 100% MD2-100(250)-145 100/250 mLPolyvinyl alcohol (PVA) 30% w/v MD2-100(250)-146 100/250 mLPolyvinylpyrrolidone (PVP) 40% w/v MD2-100(250)-21 100/250 mLPolyethyleneimine (PEI) 50% v/v MD2-100(250)-22 100/250 mLPolyacrylic acid 5100 sodium salt 50% w/v MD2-100(250)-25 100/250 mLSokalan ® CP 42 50% v/v MD2-100(250)-148 100/250 mLSokalan ® CP 5 50% w/v MD2-100(250)-149 100/250 mLSokalan ® CP 7 50% v/v MD2-100(250)-150 100/250 mLSokalan ® HP 56 40% w/v MD2-100(250)-151 100/250 mLTrimethylamine N-oxide (TMAO) 50% w/v MD2-100(250)-27 100/250 mL38 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

REAGENTSJEFFAMINES ®STOCK SOLUTIONS OF JEFFAMINES.AVAILABLE IN 100 ML OR 250 ML QUANTITIES.OptimizationORDER INFORMATIONDescription Concentration Code Available volumesJeffamine M600 - untitrated 100 mL, > pH 12 100% MD2-09 100 mLJeffamine M600 - untitrated 250 mL > pH 12 100% MD2-10 250 mLJeffamine M600, pH7 50% v/v MD2-11-100(250) 100/250 mLJeffamine D2000, pH 7 50% v/v MD2-100(250)-133 100/250 mLJeffamine ED900, pH 7 50% v/v MD2-100(250)-135 100/250 mLJeffamine ED-2001, pH7 50% v/v MD2-100(250)-23 100/250 mLJeffamine ED2003, pH 7 50% v/v MD2-100(250)-134 100/250 mLJeffamine M2005, pH 7 25% v/v MD2-100(250)-136 100/250 mLJeffamine M2005 100% MD2-100(250)-165 100/250 mLJeffamine M2070, pH 7 50% v/v MD2-100(250)-137 100/250 mLJeffamine SD2001, pH 7 50% v/v MD2-100(250)-138 100/250 mLJeffamine T403, pH 7 50% v/v MD2-100(250)-139 100/250 mLVOLATILES – ORGANICSSTOCK SOLUTIONS OF VOLATILES.AVAILABLE IN 50ML, 100 ML OR 250 ML QUANTITIES.ORDER INFORMATIONDescription Concentration Code Available volumes1,4-Butanediol 100% MD2-100(250)-61 100/250 mL1,4-Dioxane 100% MD2-100(250)-62 100/250 mL1-Propanol 100% MD2-100(250)-63 100/250 mL2-Propanol 100% MD2-100(250)-64 100/250 mLTert-butanol 100% MD2-100(250)-69 100/250 mL1,2-Propanediol 100% MD2-100(250)-98 100/250 mL1,3-Propanediol 50% v/v MD2-100(250)-99 100/250 mL1-Butanol 100% MD2-100(250)-153 100/250 mLMethanol 20% v/v MD2-100(250)-155 100/250 mLTetrahydrofuran 30% v/v MD2-100(250)-157 100/250 mLEthanol 20% v/v MD2-100(250)-158 100/250 mLTriethylammonium Phosphate 100 mM MD2-359 50 mLNorth & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com39

OptimizationREAGENTSNON-VOLATILES – ORGANICSSTOCK SOLUTIONS OF NON-VOLATILES.AVAILABLE IN 50 ML, 100 ML OR 250 ML QUANTITIES.ORDER INFORMATIONDescription Concentration Code Available volumes1,2,3-heptanetriol 0.2% w/v MD2-300 0.25 mL1,6-hexanediol 70% w/v MD2-100(250)-10 100/250 mLL-Arginine 2 M MD-100(250)-113 100/250 mLDi-ethylene glycol 50% v/v MD2-100(250)-114 100/250 mLDimethyl Sulfoxide (DMSO) 30% v/v MD2-100(250)-159 100/250 mLEthylene glycol 100% MD2-100(250)-60 100/250 mLFoscarnet (phosphoformic acid) 80 mM MD2-354 50 mLGlycerol 100% MD2-100(250)-65 100/250 mLGlycine 2 M MD2-100(250)-105 100/250 mLGlutaric Acid pH 6.0 15 mM ( 0.2%) MD2-355 50 mLL-Glutathione reduced 100 mM MD2-356 50 mLLysine HCl 2 M MD2-100(250)-106 100/250 mL2-methyl-2,4-pentanediol (MPD) 100% MD2-100(250)-24 100/250 mLPenta-ethylene glycol 0.4 M MD2-100(250)-104 100/250 mLPolyvinylpyrrolidone (PVP) 40% w/v MD2-100(250)-21 100/250 mLL-proline 2 M MD2-100(250)-107 100/250 mLSpermidine 50 mM (0.7%) MD2-357 50 mLSuccinic acid (butanedioic acid) pH 7 1.2 M MD2-100(250)-167 100/250 mLTaurine (2-aminoethanesulfonic acid) 16 mM (0.2 %) MD2-358 50 mLTetra-ethylene glycol 40% v/v MD2-100(250)-103 100/250 mLTrehalose 40% w/v MD2-100(250)-109 100/250 mLTriethylene glycol (TEG) 100% MD2-100(250)-26 100/250 mLXylitol 70% w/v MD2-100(250)-110 100/250 mLPOLYAMINESSTOCK SOLUTIONS OF POLYAMINES.AVAILABLE IN 50 ML OR 100 ML QUANTITIES.ORDER INFORMATIONDescription Concentration Code Available volumesSpermine HCl 100 mM MD2-50(100)-176 50/100 mLSpermidine 50 mM MD2-357 50 mL40 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

REAGENTSSALTSSTOCK SOLUTIONS OF SALTS AND ADDITIVES.AVAILABLE IN 100 ML OR 250 ML QUANTITIES.OptimizationORDER INFORMATIONDescription Concentration Code Available volumesAmmonium acetate 2 M MD2-100(250)-126 100/250 mLAmmonium chloride 3.5 M MD2-100(250)-29 100/250 mLAmmonium citrate tribasic 1 M MD2-100(250)-171 100/250 mLAmmonium dihydrogen phosphate 1.5 M MD2-100(250)-30 100/250 mLAmmonium formate 5 M MD2-100(250)-31 100/250 mLAmmonium hydrogen citrate -dibasic 2.5 M MD2-100(250)-32 100/250 mLAmmonium hydrogen phosphate -dibasic 2.5 M MD2-100(250)-33 100/250 mLAmmonium nitrate 5 M MD2-100(250)-34 100/250 mLAmmonium sulfate 3.5 M MD2-100(250)-35 100/250 mLBarium chloride dihydrate 1 M MD2-100(250)-36 100/250 mLCadmium chloride hemi(pentahydrate) 1 M MD2-100(250)-37 100/250 mLCadmium sulfate 8/3 hydrate 1 M MD2-100(250)-38 100/250 mLCalcium acetate hydrate 1M MD2-100(250)-39 100/250 mLCalcium chloride dihydrate 2 M MD2-100(250)-40 100/250 mLCesium chloride 1 M MD2-100(250)-41 100/250 mLChromium (III) chloride hexahydrate 0.5 M MD2-50(100)-172 50/100 mLCobalt chloride hexahydrate 1 M MD2-100(250)-42 100/250 mLCopper(II) chloride 0.1 M MD2-50(100)-173 50/100 mLHexaminecobalt(III) chloride 0.1 M MD2-100(250)-175 100/250 mLLithium acetate dihydrate 2 M MD2-100(250)-154 100/250 mLLithium chloride 5 M MD2-100(250)-43 100/250 mLLithium citrate tribasic tetrahydrate 1 M MD2-100(250)-44 100/250 mLLithium formate 2.5 M MD2-100(250)-160 100/250 mLLithium sulfate monohydrate 2 M MD2-100(250)-45 100/250 mLMagnesium acetate tetrahydrate 1 M MD2-100(250)-46 100/250 mLMagnesium chloride hexahydrate 5 M MD2-100(250)-47 100/250 mLMagnesium formate dihydrate 1 M MD2-100(250)-48 100/250 mLMagnesium nitrate hexahydrate 3 M MD2-100(250)-49 100/250 mLMagnesium sulphate heptahydrate 1 M MD2-100(250)-174 100/250 mLNickel chloride 1 M MD2-100(250)-50 100/250 mLNickel II sulfate hexahydrate 2 M MD2-100(250)-51 100/250 mLPotassium acetate 2 M MD2-100(250)-156 100/250 mLPotassium bromide 2 M MD2-100(250)-52 100/250 mLcontinued…North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com41

OptimizationREAGENTSSALTS CONT’DSTOCK SOLUTIONS OF SALTS AND ADDITIVES.AVAILABLE IN 100 ML OR 250 ML QUANTITIES.ORDER INFORMATIONDescription Concentration Code Available volumesPotassium chloride 2 M MD2-100(250)-53 100/250 mLPotassium fluoride 0.5 M MD2-342 50 mLPotassium formate 5 M MD2-100(250)-54 100/250 mLPotassium hydrogen phosphate (dibasic) 1 M MD2-100(250)-163 100/250 mLPotassium di-hydrogen phosphate (monobasic) 1 M MD2-100(250)-164 100/250 mLPotassium iodide 2 M MD2-100(250)-55 100/250 mLPotassium nitrate 2 M MD2-100(250)-56 100/250 mLPotassium silicate 0.5 M MD2-343 50 mLPotassium sodium tartrate 2 M MD2-100(250)-57 100/250 mLPotassium thiocyanate 2.5 M MD2-100(250)-58 100/250 mLRubidium chloride 0.5 M MD2-344 50 mLSodium acetate 1 M MD2-100(250)-170 100/250 mLSodium azide 0.5 M MD2-345 0.25 mLSodium bromide 2.5 M MD2-100(250)-59 100/250 mLSodium chloride 2 M MD2-100(250)-116 100/250 mLSodium citrate 1.5 M MD2-100(250)-127 100/250 mLSodium fluoride 0.8 M MD2-100(250)-86 100/250 mLSodium formate 5 M MD2-100(250)-87 100/250 mLSodium hydrogen phosphate dibasic 1 M MD2-100(250)-119 100/250 mLSodium iodide 2 M MD2-100(250)-88 100/250 mLSodium malonate 3 M MD2-100(250)-89 100/250 mLSodium metaborate 2 M MD2-100(250)-90 100/250 mLSodium nitrate 5 M MD2-100(250)-91 100/250 mLSodium oxamate 0.5 M MD2-100(250)-111 100/250 mLSodium/potassium Phosphate 0.5 M MD2-100(250)-168 100/250 mLSodium phosphate dibasic 1 M MD2-100(250)-169 100/250 mLSodium sulfate 1 M MD2-100(250)-92 100/250 mLSodium tartrate 2 M MD2-100(250)-93 100/250 mLSodium thiocyanate 2.5 M MD2-100(250)-94 100/250 mLStrontium chloride hexahydrate 0.1 M MD2-50(100)-177 50/100 mLZinc acetate 1 M MD2-100(250)-95 100/250 mLZinc chloride 2 M MD2-100(250)-96 100/250 mLZinc nitrate hexahydrate 100 mM MD2-353 50 mLZinc sulfate heptahydrate 1 M MD2-100(250)-97 100/250 mL42 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

REAGENTSOTHER REAGENTSSTOCK SOLUTIONS OF OTHER REAGENTS.AVAILABLE IN 100 ML OR 250 ML QUANTITIES.OptimizationORDER INFORMATIONDescription Concentration Code Available volumesWater, 18 MegaOhm purified/filtered MD2-69 1LConc. HCl for titration 37% MD2-100(250)-117 100 mLSodium Hydroxide solution for titration 10 M MD2-100(250)-118 100 mLNDSB-201 1M MD2-100(250)-161 100/250 mLFluorescent 'Green Screen' Dye, 20X stock 20 X MD2-13 100 mLMineral oil MD2-71 100 mLCRYOPROTECTANTSSTOCK SOLUTIONS OF CRYOPROTECTANTS.AVAILABLE IN 50 ML, 100 ML OR 250 ML QUANTITIES UNLESS SPECIFIED.ORDER INFORMATIONDescription Concentration Code Available volumesDi-ethylene glycol 50% v/v MD2-100(250)-114 100/250 mLDimethyl Sulfoxide (DMSO) 30% v/v MD2-100(250)-159 100/250 mLEthylene glycol 100% MD2-100(250)-60 100/250 mLGlycerol 100% MD2-100(250)-65 100/250 mLMaltose 30% w/v MD2-100(250)-178 100/250 mL2-methyl-2,4-pentanediol (MPD) 100% MD2-100(250)-24 100/250 mLNDSB-201 1M MD2-100(250)-161 100/250 mLParrafin Oil 100% MD2-03 100 mL1,2-propanediol 100% MD2-100(250)-98 100/250 mLTrehalose 40% w/v MD2-100(250)-109 100/250 mLXylitol 70% w/v MD2-100(250)-110 100/250 mLCryoProtX - Cryoprotectant Kit - MD1-61 46 x 1.5 mL kitNorth & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com43

OptimizationREAGENTSBUFFER STOCKSBUFFER STOCKS SOLUTIONS.AVAILABLE IN 100 ML QUANTITIES.PLEASE SPECIFY pH WHEN ORDERINGN.B. If ordering a buffer to optimise a screen condition, please specifythe exact crystal growth screen when ordering. If you have a specificrequirement for the titrant (e.g. HCl or acetic acid) please specify orcontact Molecular Dimensions to discuss.ORDER INFORMATIONDescription Concentration Code Available volumesADA 1 M MD2-001-PH 100 mLAmmonium acetate 1 M MD2-002-PH 100 mLBicine 1 M MD2-003-PH 100 mLBis Tris 1 M MD2-004-PH 100 mLBis Tris Propane 1 M MD2-005-PH 100 mLCAPS 1 M MD2-006-PH 100 mLCAPSO 0.5 M MD2-007-PH 100 mLCHES 1 M MD2-008-PH 100 mLCitrate 1 M MD2-009-PH 100 mLEPPS 1 M MD2-031-PH 100 mLGlycine 1 M MD2-010-PH 100 mLHEPES 1 M MD2-011-PH 100 mLImidazole 1 M MD2-012-PH 100 mLMES 1 M MD2-013-PH 100 mLMES/Bis Tris 0.5 M MD2-030-PH 100 mLKMES 1 M MD2-014-PH 100 mLMOPS 1 M MD2-015-PH 100 mLPhosphate/citrate 0.5 M MD2-016-PH 100 mLPIPES 1 M MD2-017-PH 100 mLPotassium citrate 1 M MD2-029-PH 100 mLPotassium phosphate 1 M MD2-018-PH 100 mLSodium acetate 1 M MD2-019-PH 100 mLSodium borate 0.1 M MD2-020-PH 100 mLSodium cacodylate 1 M MD2-021-PH 100 mLSodium citrate 1 M MD2-022-PH 100 mLSodium HEPES 1 M MD2-023-PH 100 mLSodium phosphate 0.5 M MD2-024-PH 100 mLSodium/potassium phosphate 0.5 M MD2-025-PH 100 mLTricine 1 M MD2-026-PH 100 mLTRIZMA (Tris) 1 M MD2-027-PH 100 mLTris hydrochloride 1 M MD2-028-PH 100 mL44 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

REAGENTSSPECIAL BUFFERSSPECIAL BUFFERS SOLUTIONS.AVAILABLE IN 100 ML OR 250 ML QUANTITIES.FOR DETAILS SEE PAGE 51.OptimizationORDER INFORMATIONDescription Concentration Code Available volumesSPG pH4 and pH10 1 M each MD2-59 100 mL x 2MIB pH4 and pH10 1 M each MD2-60 100 mL x 2PCTP pH4 and pH9.5 1 M each MD2-61 100 mL x 2MMT pH4 and pH9 1 M each MD2-62 100 mL x 2CHC buffer pH4 and pH10 1 M each MD2-63 100 mL x 2AAB buffer pH4 and pH9 1 M each MD2-64 100 mL x 2TBG buffer pH4 and pH9 1 M each MD2-65 100 mL x 2Imidazole / DL Malic acid 1 M each MD2-66 100 mL x 2Really Useful Buffer Kit - contains all 8 buffer systems 1 M each MD2-100 8 x 50 mL kitReally Useful Buffer kit - contains all 8 buffer systems 1 M each MD2-101 8 x 10 mL kitMORPHEUS ® MIXESSTOCK SOLUTIONS OF MORPHEUS MIXES.AVAILABLE IN 100 ML OR 250 ML QUANTITIES.ORDER INFORMATIONDescription Code Available volumesDivalents Mix MD2-100(250)-70 100/250 mLHalogens Mix MD2-100(250)-71 100/250 mLNPS Mix MD2-100(250)-72 100/250 mLAlcohols Mix MD2-100(250)-73 100/250 mLEthylene Glycols Mix MD2-100(250)-74 100/250 mLMonosaccharides Mix MD2-100(250)-75 100/250 mLCarboxylic Acids Mix MD2-100(250)-76 100/250 mLAmino Acids Mix MD2-100(250)-77 100/250 mLP550MME_P20K Mix MD2-100(250)-81 100/250 mLEDO_P8K Mix MD2-100(250)-82 100/250 mLGOL_P4K Mix MD2-100(250)-83 100/250 mLMPD_P1KP3350 Mix MD2-100(250)-84 100/250 mLBuffer System 1 MD2-100(250)-100 100/250 mLBuffer System 2 MD2-100(250)-101 100/250 mLBuffer System 3 MD2-100(250) -102 100/250 mLNorth & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com45

OptimizationREAGENTSMORPHEUS ® INDIVIDUALSTOCK REAGENTSBUFFER STOCKS SOLUTIONS.AVAILABLE IN 100 ML OR 250 ML QUANTITIES.ORDER INFORMATIONDescription Concentration Code Available volumesMagnesium chloride hexahydrate 5 M MD2-100(250)-47 100/250 mLCalcium chloride dihydrate 2 M MD2-100(250)-40 100/250 mLSodium bromide 2.5 M MD2-100(250)-59 100/250 mLSodium fluoride 0.8 M MD2-100(250)-86 100/250 mLSodium iodide 2 M MD2-100(250)-88 100/250 mLAmmonium sulfate 3.5 M MD2-100(250)-35 100/250 mLSodium nitrate 5 M MD2-100(250)-91 100/250 mLSodium Hydrogen Phosphate dibasic 1 M MD2-100(250)-119 100/250 mL1,6-hexanediol 70% w/v MD2-100(250)-10 100/250 mL1,2-propanediol 100% MD2-100(250)-98 100/250 mL1,3-Propanediol 50% v/v MD2-100(250)-99 100/250 mL1-Butanol 100% MD2-100(250)-153 100/250 mL1,4-Butanediol 100% MD2-100(250)-61 100/250 mL2-Propanol 100% MD2-100(250)-64 100/250 mLTetra-ethylene glycol 40% v/v MD2-100(250)-103 100/250 mLPenta-ethylene glycol 0.4 M MD2-100(250)-104 100/250 mLDi-ethylene glycol 50% v/v MD2-100(250)-114 100/250 mLTriethylene glycol (TEG) 100% MD2-100(250)-26 100/250 mLN-acetyl-D-glucosamine 1 M MD2-100(250)-120 100/250 mLL-Fucose 1 M MD2-100(250)-121 100/250 mLD-Galactose 1 M MD2-100(250)-122 100/250 mLD-Glucose 1 M MD2-100(250)-123 100/250 mL46 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

REAGENTSMORPHEUS ® INDIVIDUALSTOCK REAGENTS CONT’DOptimizationBUFFER STOCKS SOLUTIONS.AVAILABLE IN 100 ML OR 250 ML QUANTITIES.ORDER INFORMATIONDescription Concentration Code Available volumesD-Mannose 1 M MD2-100(250)-124 100/250 mLD-Xylose 1 M MD2-100(250)-125 100/250 mLSodium oxamate 0.5 M MD2-100(250)-111 100/250 mLSodium formate 5 M MD2-100(250)-87 100/250 mLPotassium sodium tartrate 2 M MD2-100(250)-57 100/250 mLAmmonium Acetate 2 M MD2-100(250)-126 100/250 mLSodium Citrate 1.5 M MD2-100(250)-127 100/250 mLGlycine 2 M MD2-100(250)-105 100/250 mLLysine HCl 2 M MD2-100(250)-106 100/250 mLDL-Alanine 2 M MD2-100(250)-112 100/250 mLDL-Glutamic acid 2 M MD2-100(250)-128 100/250 mLDL-serine 2 M MD2-100(250)-129 100/250 mLPEG 500 MME 50% v/v MD2-100(250)-66 100/250 mLPEG 20,000 50% w/v MD2-100(250)-16 100/250 mLEthylene glycol 100% MD2-100(250)-60 100/250 mLPEG 8000 50% w/v MD2-100(250)-13 100/250 mLGlycerol 100% MD2-100(250)-65 100/250 mLPEG 4000 50% w/v MD2-100(250)-11 100/250 mL2-methyl-2,4-pentanediol (MPD) 100% MD2-100(250)-24 100/250 mLPEG 1000 50% w/v MD2-100(250)-5 100/250 mLPEG 3350 50% w/v MD2-100(250)-9 100/250 mLMorpheus Additive OptiMax Kit* MD1-58 43 x 10 mL kit* Morpheus Additive OptiMax Kit contains 43 x 10 mL of each of the above stock reagents.North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com47

OptimizationREAGENTSMIDAS REAGENTSSTOCK SOLUTIONS OF MIDAS REAGENTS.AVAILABLE IN 50 ML, 100 ML OR 250 ML QUANTITIES.ORDER INFORMATIONDescription Concentration Code Available volumesAcrylic acid/maleic acid copolymer (50:50), Na salt 35% v/v MD2-100(250)-130 100/250 mLGlycerol ethoxylate 50% v/v MD2-100(250)-132 100/250 mLJeffamine D2000, pH 7 50% v/v MD2-100(250)-133 100/250 mLJeffamine ED2003, pH 7 50% v/v MD2-100(250)-134 100/250 mLJeffamine ED900, pH 7 50% v/v MD2-100(250)-135 100/250 mLJeffamine M2005 100% MD2-100(250)-165 100/250 mLJeffamine M2070, pH 7 50% v/v MD2-100(250)-137 100/250 mLJeffamine SD2001, pH 7 50% v/v MD2-100(250)-138 100/250 mLJeffamine T403, pH 7 50% v/v MD2-100(250)-139 100/250 mLJeffamine M600, pH7 50% v/v MD2-11-100(250) 100/250 mLPentaerythritol ethoxylate (15/4 EO/OH) 50% v/v MD2-100(250)-140 100/250 mLPentaerythritol ethoxylate (3/4 EO/OH) 50% v/v MD2-100(250)-141 100/250 mLPentaerythritol propoxylate (5/4 PO/OH) 50% v/v MD2-100(250)-115 100/250 mLPentaerythritol propoxylate (17/8 PO/OH) 50% v/v MD2-100(250)-142 100/250 mLPolyethyleneimine (PEI) 50% v/v MD2-100(250)-22 100/250 mLPolyacrylate 2100, sodium salt 50% w/v MD2-100(250)-143 100/250 mLPolyacrylate 5100, sodium salt 50% w/v MD2-100(250)-144 100/250 mLPolyvinylpyrrolidone (PVP) 40% w/v MD2-100(250)-21 100/250 mLPolypropylene glycol 400 100% MD2-100(250)-145 100/250 mLPolyvinyl alcohol (PVA) 30% w/v MD2-100(250)-146 100/250 mLSokalan ® CP 5 50% w/v MD2-100(250)-149 100/250 mLSokalan ® CP 7 50% v/v MD2-100(250)-150 100/250 mLSokalan ® HP 56 40% w/v MD2-100(250)-151 100/250 mLMIDAS OptiMax Kit - MD1-62 27 x 10 mL kit48 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

REAGENTSMEMADVANTAGE REAGENTSSTOCK SOLUTIONS OF MEMADVANTAGE REAGENTS.AVAILABLE IN QUANTITIES LISTED BELOW.OptimizationORDER INFORMATIONDescription Concentration Code Available volumes1,2,3-heptanetriol 0.2% w/v MD2-300 0.25 mLBenzamidine HCl 20% w/v MD2-301 0.25 mLEDTA 100 mM MD2-302 50 mLEGTA 10 mM MD2-303 50 mLAnzergent 3-12 30 mM MD2-304 0.25 mLC-Hega-11 11.5 mM MD2-305 0.25 mLCHAPS 60 mM MD2-306 0.25 mLCymal 1 340 mM MD2-307 0.25 mLCymal 2 120 mM MD2-308 0.25 mLCymal 4 76 mM MD2-309 0.25 mLCymal 5 37 mM MD2-310 0.25 mLCymal 6 5.6mM MD2-311 0.25 mLCymal 7 1.9 mM MD2-312 0.25 mLFos-Choline 9 39.5 mM MD2-313 0.25 mLDDM (n-dodecyl-β-D-maltopyranoside) 1.7 mM MD2-314 0.25 mLSucrose monodecanoate (sucrose monocaprate) 25 mM MD2-315 0.25 mLDeoxy-BigCHAP 14 mM MD2-316 0.25 mLDM (n-decyl-β-D-maltopyranoside) 18 mM MD2-317 0.25 mLDSM (n-decyl-β-d-thiomaltopyranoside) 9 mM MD2-318 0.25 mLDNG (decyl maltose neopentyl glycol) 3.6 mM MD2-319 0.25 mLFos-Choline 12 15 mM MD2-320 0.25 mLHEGA 11 14 mM MD2-321 0.25 mLHEGA 10 70 mM MD2-322 0.25 mLLDAO (n-dodecyl-N,N-dimethylamine-N-oxide) 15 mM MD2-323 0.25 mLLNG (lauryl maltose neopentyl glycol) 1 mM MD2-324 0.25 mLMERPOL HCS 5% w/v MD2-325 0.25 mLNG (n-nonyl-β-D-glycopyranoside) 65 mM MD2-326 0.25 mLNM (n-nonyl-β-D-maltopyranoside) 60 mM MD2-327 0.25 mLOG (n-octyl-β-D-glucoside) 190 mM MD2-328 0.25 mLOM-fluorinated (octyl maltoside flourinated) 10.2 mM MD2-329 0.25 mLONG (octyl glucose neopentyl glycol) 10.2 mM MD2-330 0.25 mLTri DM (n-tridecyl-β-D-maltoside) 0.3 mM MD2-331 0.25 mLTripao 45 mM MD2-332 0.25 mLcontinued…North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com49

OptimizationREAGENTSMEMADVANTAGE REAGENTS CONT’DSTOCK SOLUTIONS OF MEMADVANTAGE REAGENTS.AVAILABLE IN QUANTITIES LISTED BELOW.ORDER INFORMATIONDescription Concentration Code Available volumesUDM (n-undecyl-β-D-maltoside) 5.9 mM MD2-333 0.25 mLC12E8 (octaethylene glycol monododecyl ether) 0.9 mM MD2-334 0.25 mLC8E4 (tetraethylene glycol monooctyl ether) 80 mM MD2-335 0.25 mLDMG (n-dodecyl-N,N-dimethylglycine) 15 mM MD2-336 0.25 mLHTG (n-heptyl-β-D-thioglucopyranoside) 290 mM MD2-337 0.25 mLUDTM (n-undecyl-B-D-thiomaltopyranoside) 2.1 mM MD2-338 0.25 mLLAPAO (3-laurylamido-N,N'-dimethylpropyl amino oxide 15.6 mM MD2-339 0.25 mLDeuterium Oxide 1 mM MD2-340 0.25 mLGly-Gly-Gly 300 mM MD2-341 50 mLPotassium fluoride 0.5 M MD2-342 50 mLPotassium Silicate 0.5 M MD2-343 50 mLRubidium Chloride 0.5 M MD2-344 50 mLSodium Azide 0.5 M MD2-345 0.25 mLGadolinium(III) Chloride hexahydrate 100 mM MD2-348 1 mLManganese(II) chloride tetrahydrate 100 mM MD2-349 50 mLOsmium(III) Chloride hydrate 100 mM MD2-350 1 mLSamarium(III) Chloride 100 mM MD2-351 50 mLStrontium Chloride hexahydrate 100 mM MD2-352 50 mLZinc Nitrate hexahydrate 100 mM MD2-353 50 mLFoscarnet (phosphoformic acid) 80 mM MD2-354 50 mLGlutaric Acid pH 6.0 15 mM (0.2%) MD2-355 50 mLL-Glutathione reduced 100 mM MD2-356 50 mLSpermidine 50 mM (0.7%) MD2-357 50 mLTaurine 16 mM (0.2%) MD2-358 50 mLTriethylammonium Phosphate 100 mM MD2-359 50 mL2-Mercaptoethanol 30 mM MD2-360 0.25 mLDTT 100 mM MD2-361 0.25 mLTCEP 100 mM MD2-362 0.25 mLSucrose monododecanoate 3 mM MD2-363 0.25 mL50 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

REAGENTSTHE REALLY USEFULBUFFER KIT 8 BROAD-RANGE BUFFERING SYSTEMS. ALLOWUN-COUPLING OF THE CHEMICAL NATURE OF THEBUFFERING COMPONENTS FROM pH.REAGENTSGELLED-SURFACE KITGELLED-SURFACE CRYSTALLIZATION IS AN IMPROVEDVARIATION OF THE CONTAINERLESS CRYSTALLIZATIONMETHOD 1 WHERE THE DENSE, LOWER AND EXPENSIVEFLUOROSILICON OIL IS REPLACED WITH AN INERT (NON-AQUEOUS) GELLED LAYER, THUS PROVIDING A STABLEINTERFACE TO THE UPPER LAYER OIL.OptimizationIt is not unusual to have specific interactions between buffers andprotein molecules. An extensive list of buffers can be found instructures deposited in the PDB. Thus, a dilemma arises duringoptimization of how to uncouple the chemical nature and thebuffering properties of the `buffer' component of a crystallizationcocktail. To address this challenge, Janet Newman (Newman,J.,2004) has developed a series of buffers by mixing differentbuffering components. This set of buffers, complimented by theimidazole-malate buffer from our Stura and Macrosol Screens, isnow presented as a kit.Features of the Really Useful Buffer Kit:n Broad-range buffer systems 1 - 4 .n Linear response to pH on mixing. (not imidazole/malate)n Un-couple the chemical nature of the buffers from pH control.n Individual broad-range buffers (100 mL) also available to order.ORDER INFORMATIONDescription Pack CodeReally Useful Buffer Kit 10 mL each MD2-101Really Useful Buffer Kit 50 mL each MD2-100SPGSuccinic acid, Sodiumdihydrogen phosphate,Glycine (pH 4) & (pH 10) MD2-59MIBSodium malonate, Imidazole,Boric acid (pH 4) & (pH 10) MD2-60PCTPSodium propionate,Sodium cacodylate,Bis-Tris propane (pH 4)& (pH 9.5)MD2-61MMTCHCAABTBGImidazole / malateL-Malic acid, MES, Tris(pH 4) & (pH 9)Citric acid, HEPES, CHES(pH 4) & (pH 10)Sodium acetate, ADA,Bicine(pH 4) & (pH 9)Sodium tartrate, Bis Tris,Glycylglycine (pH 4)& (pH 9)MD2-62MD2-63MD2-64MD2-65Imidazole / DL-Malic acid MD2-66Features of Gelled-Surface Crystallization:n Promote the growth of fewer crystals.n Promote the growth of larger crystals.n Avoid attachment of crystals to the container surface.ORDER INFORMATIONDescription Pack CodeGelled-Surface Kit5 x 96-wellplates plusupper layer oil MD1-12MICROBATCH OILSWHILST NOT STRICTLY OPTIMIZATION, THE MICROBATCH 2,3TECHNIQUE UNDER OIL HAS EVOLVED FURTHER WITH THEUSE OF MIXED OILS 4 (AL’S OILS) TO ALLOW A DEGREE OFEVAPORATION THROUGH THE OIL LAYER THUS MIMICKINGVAPOUR DIFFUSION.n Molecular Dimensions supplies both paraffin and silicon oil.ORDER INFORMATIONDescription Pack CodeContainerless Crystallization Oils 2 x 50 mL MD2-05Paraffin Oil 100 mL MD2-03Silicon Oil 100 mL MD2-04Volatile Oil 100 mL MD2-06References1. Berman H. M., Westbrook J., Feng Z., Gilliland G., Bhat T. N., Weissig H., Shindyalov I. N. &Bourne P. E. (2000). Nucleic Acids Res. 28, 235- 242.2. Jancarik J. & Kim S.-H. (1991). J. Appl. Cryst. 24, 409-411.3. Kleywegt G. J. & Jones T. A. (1998). Acta Cryst. D54, 1119-1131.4. Newman J. (2004) Acta Cryst. D60, 610-612.References1. Chayen et al Appl. Cryst. (1990) 23, 297.2. D’Arcy et al J. of Cryst.Growth(1996) 168, 175-180.3. Lorber B. and Giegé R. J. of Cryst. Growth (1996) 168, 204-215.4. Chayen N.E. Protein Engineering (1996) 9, 10, 927-929.5. Allan D'Arcy et al. Acta Cryst. (2003). D59, 396-399.North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com51

ADVANCEDMETHODS INPROTEINCRYSTALLIZATIONNové Hrady 2012I have written down some of the methods in the form ofexercises, to give you step by step protocols of settingup the experiments.The methods are mostly for optimization – in cases thatyou get poor crystals that need improvement. Some canalso be applied for screening as mentioned in thespecific protocols.These methods have been successful in yieldingdiffraction quality crystals of a variety of proteins in caseswhere conventional methodology failed. I have usedlysozyme and trypsin as models in some of the protocolsin order to have examples and results that you can seewithin a short time, but of course, the aim is to use thesetechniques with your problem proteins.From left: Dr Lata Govada, Prof Naomi Chayen and Dr Sahir Khurshid.Practical ExercisesProf Naomi ChayenImperial College LondonThere is no magic bullet to solve all the crystallizationproblems however having a portfolio of differenttechniques is very helpful.GOOD LUCK!NaomiFirst published at the FEBS Practical Course –Advanced methods on macromolecular crystallization.Nové Hrady, Czech Republic 2012.52 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

EXERCISE 1:Insertion of oil barrier to slow downvapor diffusion experimentsChayen N.E. (1997) J. Appl. Cryst. 30, 198-202The purpose of this experiment is to improve the size andquality of crystals which are obtained as showers of small uselesscrystals, twinned crystals or precipitate. The experiment alsodemonstrates the effect that oil-over-the reservoir has on therate of equilibration (and subsequent effects on crystal growth.)Materials:• Crystallization plates for hanging or sitting drops• cover slips• grease or sealing tape• silicone oil• paraffin oil• Protein solution• Crystallization reagentsMethod for Preparation of the oils1. Mix paraffin and silicone oils in equal volumes.2. Shake well and allow to stand for several minutes. The oilsare totally miscible once the bubbles have disappeared.Method for setting up hanging drops1. Use a XRL plate for hanging drops.2. Coat the lips of the reservoirs with grease or oil.3. Pipette 0.6 - l mL of the reservoir solution which gives you theshowers of crystals into each well.4. Pipette volumes ranging from 0.1 mL to 0.5 mL of a mixtureof paraffin and silicone over all the reservoirs except for onereservoir. The oil will form a layer above the reservoirs (figure).5. Dispense the hanging drops on the coverslips as usual bymixing the protein solution with the reservoir solution. Usethe reservoir without oil as your source of precipitant for allthe drops.6. Invert the cover slips and place over the wells containing theoil layer.7. Place the last drop over the reservoir without the oil. Thisdrop will act as your control.8. Incubate at the temperature of your choice.9. If the quality of the crystals is not sufficiently improved,repeat the protocol using different ratios of paraffin andsilicon.Method for sitting and sandwich dropsIn the case of sitting and sandwich drops, set up the trials as youwould normally do and place the layer of oil above the reservoirbefore sealing the plates with tape.Warning: This technique does not work with PEG or MPDconcentration above 13% but is very effective at concentrationsbelow 13% and at all concentrations of all salts.CrystallizationdropMixture ofparaffin &silicone oilsReservoirExpected results:Wait patiently for the results because in trials containing an oilbarrier, crystals require longer periods (e.g. 8-10 days comparedto 12-24 hours) to grow to full size, but their quality is improved.North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com53

EXERCISE 2:Setting up microbatch trialsChayen et al (1992) J.Cryst. Growth 122, 176-180.D'Arcy et al (1996) J.Cryst. Growth 168, 175-180Chayen N.E. (1997) Structure 5, 1269 - 1274.Purpose of experiment:To employ a method of crystallization with different inherentqualities to that of diffusion methods. The method is used forscreening and for optimisation.Materials required:1. Three microbatch plates.2. Low density oils:• Paraffin oil• Silicone oilProteins and buffers required:1. Lysozyme at 40 mg/mL in 10 mM sodium acetate buffer pH4.72. Precipitant: 12% NaCl in 10 mM sodium acetate buffer pH 4.7General procedure forsetting up microbatch manually:1. Pipette or dispense 5 mL of paraffin oil into a microbatchplate. The oil will spread over the plate and cover the wells.2. Using a Gilson P2 or similar pipette, withdraw 1 µl of theprecipitant solution.3. Insert the tip into the well under the surface of the oil anddispense the 1 µl drop. If you find it difficult to hold the tip inmid-oil, you can rest the edge of the tip on the floor of theplate as you dispense. As you withdraw the tip from the oil,the drop will detach from it and fall to the bottom of the well.4. Now add in the same way 1 µl of protein solution to that welland mix gently with the pipette tip. The two (separate) 1 µldrops join and become a 2 µl drop.5. Incubate at the temperature of your choice.6. Observe trials regularly under a light microscope.Microbatch is mechanically the simplest crystallization methodand is therefore increasingly applied for high throughput trialsespecially for screening.Example with lysozyme:1. Fill the microbatch dish with 5-6 mL of paraffin oil.2. Lysozyme: Make 3 rows of drops. In the first row, the drop sizewill be 0.5 µl 40 mg/mL lysozyme + 0.5 µl 12% NaCl.3. In the second row, repeat, with 1 µl + 1 µl.4. In the third row, repeat with 4 µl + 4 µl.5. Repeat the experiment in two identical dishes, one with 5 mLof silicone oil, one with 5 mL of a 50:50 mixture ofparafin:silicone and another with 5 mL of paraffin oil.Tip: Instead of mixing the precipitant (NaCl) with the protein inthe plate as described above in the general procedure, you canalso pre-mix the two ingredients in an eppendorf tube then addthe ready made drops of 1 µl, 2 µl and 8 µl respectively to theplate.Expected results:The lysozyme crystals will appear over 2 to 7 days. See if the sizeof crystals varies as a function of drop size in the lysozymeexperiment. The silicone oil should give faster results andparaffin oil the slowest.CrystallisationdropLow densityoil54Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

Method for setting up using a robot:There are several robots for setting up screening experiments inmicrobatch. The precipitant solutions are transferredsimultaneously from stock screening solutions to crystallizationplates by any number of syringes depending on the roboticsystem. The drops are dispensed under oil and the protein isadded to the precipitant drops using a dedicated syringe for theprotein solution either simultaneously with the screeningsolutions or at a later stage. Some of the robots have a routine ofmixing the drops.Tip: For screening purposes it is preferable to use silicone oil or amixture of paraffin and silicone oils. This allows some evaporationof the drops leading to a higher number of 'hits' and fasterformation of crystals compared to trials which are set underparaffin oil. In the case of optimization, where the conditionsneed to be known and stable, the trials must be covered byparaffin oil.N.B. Microbatch can be used for almost all the knownprecipitants, buffers and additives including detergents. The oilsdo not interfere with the common precipitants such as salts,polyethylene glycols (PEG), jeffamine MPD and even glycerol andethanol. Microbatch, though, can not be used for crystallizationtrials containing small volatile organic molecules such as dioxane,phenol, or thymol since these molecules dissolve into the oil.Crystallization of membraneproteins in microbatchChayen, N.E. and Saridakis, E. (2008) Nature Methods, 5, 147-153.An increasing number of membrane proteins in a variety ofdifferent detergents have been crystallised in microbatch underoil. Some of these had failed to crystallise by all methods otherthan microbatch. Dispensing is quick and simple even whenperformed manually and the drops in oil do not spread out asthey do in vapour diffusion over the siliconised coverslips. Usingrobots thousands of microbatch trials can be dispensed in highthroughput mode in nanolitre volumes.The microbatch can be used for both screening and foroptimisation of membrane proteins. The protocol for setting upmicrobatch experiments containing membrane proteins isidentical to that described in Exercise 2.EXERCISE 3:Harvesting and mounting crystalsfrom microbatchChayen N.E. (1998) Acta Cryst D54, 8-15Harvesting crystals from microbatch is slightly more difficult thanharvesting from coverslips or from standard sitting drops. Howeverafter some practice it can be achieved easily.Two alternative ways of harvesting crystalsfrom microbatch:Materials required:Cryoprotectant solutionPrecipitate solution at ~ 5 % higher concentration than that in thedropsMicro toolsStandard pipetteScalpelLoopsDepression platesMethod:1. Add a few microlitres of cryo-protectant solution to the dropcontaining the crystals.2. After several minutes check that the crystals are not cracked ordissolved by looking at them under the microscope. If theycrack/dissolve, adjust the concentration of cryo-protectant orchange cryo-protectant.3. Take the crystals directly out of the oil using a loop and freeze.If the above protocol proves tricky, harvest inthe following way:1. Add harvest solution (of ~5% higher concentration of precipitantthan that in the drop) into the well containingthe crystals. If you have a 1 µl drop, add 5-10 µl of harvest solution.2. Wait a short while (up to 15 minutes) to allow the crystalsto equilibrate.3. Withdraw the enlarged drop using a standard 10-100 µl pipettewhich had its tip cut off with a scalpel in order to widen its bore.4. If the crystals stick to the vessel, loosen them gently inside thedrop using micro tools or very thin strips of filter paper (the edgeof the strip that will touch the crystal is best torn rather than‘cleanly’ cut with scissors).5. Transfer the drop into a depression well containing more harvestsolution.6. From this stage onwards, handle the crystals as you wouldfrom a standard diffusion trial.North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com55

EXERCISE 4:Containerless crystallizationChayen N.E. (1996) Protein Engineering 9, 927-929.Chayen N.E. and Saridakis E. (2002) Lead Article ActaCryst. D. 58, 921-927Purpose of this experiment:(1) to aid harvesting (2) to reduce the amount of nucleation in atrial by eliminating the effects of surface contact between thecrystallization trial and its supporting vessel.Materials required:• "Gelled Surface" kit MDI-12• Paraffin oil• Lysozyme 40 mg/mL in Na acetate, pH 4.7, 12% NaClfrom the previous experimentsProcedure:• Pipette the paraffin oil into the wells of the Gelled-Surfaceplate.• Prepare the protein/precipitant drops as described formicrobatch experiments.• Insert the pipette tip into the well, under the surface of theparaffin oil, and dispense the drop.• Experiment with different size drops e.g. 1-20 µl.You can also try dispensing a drop under oil (figure a) andcompare with a drop dispended between two oils of differentdensities (Figure b).Try the method using your problem proteins that are giving youmany small unusable crystals.EXERCISE 5:Filtration experimentsChayen N.E., Saridakis E., Sear, R.P. (2006). PNAS 103: 597-601Chayen NE (2009) J. Appl. Cryst. 42, 743–744.Purpose of this experiment:Filtration will remove particles (dust, protein aggregates, fungi,bacteria, etc) and thus reduce the amount of nucleation in the trialsleading to the growth of fewer larger crystals.Materials required:• Centrifugal filters: 0.2 µm 0.1µmhttp://www.millipore.com/catalogue/item/ufc30vv25 orhttp://www.millipore.com/catalogue/item/ufc30vv00• Bench centrifuge.• 40 mg/mL lysozyme stock• 12% NaClProcedure:1. Mix 150 µl of lysozyme at 40 mg/mL with 150 µl precipitant (12%NaCl in 10 mM acetate buffer pH 4.7) in a microcentrifuge tube.2. Divide the mixture into 3 aliquots of 100 µl each:(a) leave one aliquot unfiltered(b) filter the second aliquot through a 0.22 µm filter(c) filter the third aliquot through a 0.1 µm filterFiltration is performed by placing the filters in a bench centrifugeat 2-9 g for 1 – 2 minutes.3. Dispense droplets of each aliquot for crystallization under the oil.Expected results:You will get a number of crystals in the unfiltered solution, a smallernumber of larger crystals in the solution which was filtered with a0.22 µm filter, and very few or no crystals at all in the solution whichwas filtered through the 0.1 µm filter.Warning:• You need to filter a minimum of 40 µL in order not to loseany protein.• Do not use the filtration method if your protein is sticky.56Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

EXERCISE 6:Improving crystal quality by separatingnucleation and growth in hanging drops(not using oils!)Saridakis, E. and Chayen, N.E. (2003) Biophys J. 84, 1218-1222.Chayen, N.E. (2004) Current Opinion in Structural Biology 14, 577-583.Chayen, N.E. (2005) Progress in Biophysics and MolecularBiology 88, 329-337.This is a method to get nucleation "going" and then "back off"before the nucleation becomes excessive. It can be used foroptimisation when small useless crystals are formed, andimprovement can not be obtained neither by fine-tuning theconcentrations of the protein and precipitating agents nor byaddition of additives. In practice, cover-slips holding the drops areincubated for some time over reservoir solutions that normally givemany small crystals and after given times the cover-slips aretransferred over reservoirs with lower precipitant concentrationsthat would normally yield clear drops. This method can also beused for screening.Example for optimisation:Materials needed:Trypsin from porcine pancreas(Sigma catalogue no: T-0134)dissolved in de-ionised water at a concentration of 40 mg/mL.1 M Tris pH 8.43M ammonium sulphateMineral oil (viscosity of ~5) to put round the edge of the XRL platesfor sealing the cover-slips (not the thick Vaseline usually applied asa seal).Standard XRL plates and siliconised cover-slips .Pasteur pipette with rubber squeezer or plastic squeezer pipettes.Procedure:1) Set up 6 trials under conditions that would give you low qualitycrystals, in this example it is reservoir solutions containing 1.7Mammonium sulphate and 0.1M Tris pH 8.4. Dispense those into6 wells of a plate.2) Prepare 6 reservoirs with solutions containing precipitantconcentration that would result in producing a clear drop ifcrystallization drops were set up and left to incubate underthese conditions. In this example it is reservoir solutionscontaining 1.3M ammonium sulphate and 0.1M Tris pH 8.4.Dispense those into 6 wells of another row in the same plate.3) Set up hanging drops by taking equal volumes (recommended1 microlitre + 1 microlitre) of the 40mg/mL protein solution andmixing it with the well solutions containing the 1.7 Mammonium sulphate.4) After 2 hours transfer one of the cover-slips from over the wellscontaining the 1.7M ammonium sulphate and just place it overone of the wells which contain 1.3M ammonium Sulphate(Figure). The transfer should only take 1-2 seconds.5) Repeat the same with the other drops after 3, 4 and 6 hours.6) Leave 2 drops at the high concentration (1.7M) as controls7) set two drops at the low concentration (1.3M) as controls.8) Observe the results each day for two weeks.Expected results:The drops which are left over the reservoirs containing 1.7 Mammonum sulphate will have clusters of crystals overnight.The drops hung over the 1.3M should be clear. The dropstransferred after 4 or 6 hours should produce large single crystalsafter 8-14 days.Tip: The time of transfer is selected by reference to the time inwhich it took to see the first crystals in the trials producing thepoor crystals. In the case of trypsin, crystals appeared within24 hours meaning that nucleation would have occurred anytimebetween set up of the experiments to several hours before thecrystals appeared. Hence transfer was done at intervals of 2 hoursafter set up. Trials that are transferred too soon will produce cleardrops while those that are transferred too late will yield lowquality crystals. In cases of other proteins where crystals take afew days to appear, the transfers should be performed at longertime intervals e.g. every 12 hours or so.Reservoir3 hrsCrystallizationdropReservoir withlower pp1 concentrationCrystallizationdropFor screening:3D structure screen (MD1-13)This screen consists of two sets of solutions: 24 contain sparsematrix screening conditions; the other 24 are a 70% dilution of theprecipitants in those solutions (buffer and additive concentrationsremain the same). Using this screen, the hanging drops are firstincubated for 3-6h over the solutions at high concentrations.The cover-slips holding the drops are then transferred over thereservoirs at 70% dilution.North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com 57

Introducing nucleants intocrystallization trialsChayen N.E., Saridakis E., Sear, R.P. (2006) PNAS 103, 597-601Saridakis, E. and Chayen, N.E. (2009) Trends in Biotechnology 27,99-106.Chayen, N.E. and Saridakis, E. (2008) Nature Methods, 5, 147-153.Saridakis et al. (2011) PNAS 108, 11081-11086.EXERCISE 7:What to do when you get no crystals?Khurshid, S., Govada, L. and Chayen, N.E. (2007) Crystal Growth& Design 7, 2171-2175.This is a screening technique used in vapour diffusion hangingdrops when trials remain persistently clear.Materials required:1. XRL plate, siliconised coverslips and sealing grease2. Screens of your choice3. Your protein solution1. Set up your screening as you would normally do2. Loosen the coverslips of drops that remained clear after2 weeks. The angle allows variable amounts of evaporationwithout exposing the drops3. Observe the drops every 2 hours in the first day and re-sealthe caps when the drops visibly shrink.Expected results:By loosening the coverslip you are allowing evaporation for alimited time. What you are effectively doing is inducingnucleation which you then arrest when re-sealing. This will drivethe system into supersaturation and hopefully give you a hitwhich you can then proceed to optimise using the technique ofyour choice.First, determine the metastable zone as follows:1. Note the conditions in a ‘hit’ of a screen that have yielded poorcrystals or a crystalline precipitate.2. Dispense (manually or by a robot) 10-24 trials using a crystallizationmethod of your choice, varying the protein and precipitantconcentrations in steps (as instructed in point 3 below) on a matrixgrid.3. The concentrations of protein and precipitating agents should beabove and below the concentrations that gave the conditions ofthe hit. Alternatively the pH, temperature, or another parameter towhich solubility is sensitive can be varied.4. Plot the results and you will obtain the supersolubility curve. e.g.diagram below.: For example, if a screen produced ‘hits’ atconditions of 10-22 mg/mL of protein and 1-1.5 M Ammoniumsulphate, the experiments are set at concentrations ranging from5-25 mg/mL protein versus 0.5-2.2 M ammonium sulphate, thuscovering a range of conditions above and below the ‘hits. The areajust below the curve is the metastable zone. X represent conditionsobtained from screening ‘hits’, giving low quality crystals orcrystalline precipitate. △ represent clear drops; ▲ representprecipitate.[protein] mg/ml252015105XXXsupersolubilitycurveThe method can also be used for optimization as described inGovada, L. and Chayen, N.E. (2009) Crystal Growth & Design 9,1729-1732.00.51.0 1.5 2.0 2.5[Ammonium sulphate] MInsert your nucleant at conditions just below the curve (where theheads of the arrows are pointing) using fine tweezers and let the trialincubate as you would normally do.Nucleant are used mostly for optimization but can also be used forscreening. When nucleants are placed in supersaturated conditions,crystals will usually appear faster.58Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

References on crystallization methodsin microbatch and vapour diffusion(and some in microgravity)Chayen, Shaw Stewart, Maeder & Blow (1990). An automated system for microbatch proteincrystallisation and screening. J. Appl. Cryst. 23, 297-302.Chayen, Shaw Stewart & Blow (1992). Microbatch crystallisation under oil - a new techniqueallowing many small-volume crystallisation trials. J. Crystal Growth, 122, 176-180.Chayen, Shaw Stewart & Baldock (1994). New developments of the IMPAX small-volumecrystallisation system. Acta Cryst. D . 50, 456-458.Chayen, Radcliffe & Blow (1993). Control of nucleation in the crystallisation of lysozyme. ProteinScience, 2, 113-118.Normile (1995). Search for better crystals in inner and outer space. Science 270, 1921-1922.Baldock, Mills & Shaw Stewart (1996). A comparison of microbatch and vapour diffusion for initialscreening of crystallization conditions. J. Crystal Growth, 168, 170-174..D’Arcy, Elmore, Stihle & Johnston (1996). A novel approach to crystallising proteins under oil. J.Crystal Growth, 168, 175-180.Chayen (1996). A novel technique for containerless protein crystallisation. Protein Engineering 9,No. 927-929.Lorber and Giege (1996). Containerless protein crystallisation in floating drops. J. Cryst. Growth,168, 204-215.Chayen et al. (1996). Trends and challanges in experimental macromolecular crystallography. Q.Rev. Biophys. 29, 227-278.Chayen (1997). A novel technique to control the rate of vapour diffusion, giving larger proteincrystals. J. Appl. Cryst. 30, 198-202.Chayen (1997) The role of oil in macromolecular crystallization. Structure 5, 1269-1274.Chayen (1998) Comparative studies of protein crystallisation by vapour diffusion and microbatch.Acta Cryst. D54, 8-15.Chayen, N.E. and Helliwell, J.R., “Protein Crystallography: the Human Genome in 3 - D” PhysicsWorld 11 (1998), 43-48.Chayen, N.E., “Novel Methodology in the Crystallisation of Biological Macromolecules” J. Cryst.Growth 198/199 (1999) 649-655.Chayen, N.E. and Helliwell, J.R., “Space-grown Crystals May Prove their Worth” Nature 398,(1999) 20.Chayen, N.E. “New Trends in Macromolecular Crystallisation” Research Trends (1999) 5, 141-149.Saridakis, E. and Chayen, N.E. “Improving protein crystal quality by de coupling nucleation andgrowth in vapour diffusion”. Protein Science (2000), 9 755-757.Jovine L. (2000) A simple technique to control macromolecular crystal nucleation efficiently usinga standard vapour-diffusion set up J. Appl. Cryst. 33, 988-989.Chayen, N.E. and Saridakis E. (2001) Is Lysozyme the ideal model protein? J. Cryst. Growth 232,262-264.Chayen, N.E., Saridakis, E., El-Bahar, R. and Nemirovsky, Y. (2001) Porous silicon: an effectivenucleation-inducing material for protein crystallisation. J. Mol Biol. 312, 591-595.Moreno, A., Saridakis, E. and Chayen, N.E. (2002) Combination of oils and gels for enhancing thegrowth of protein crystals. J Appl Cryst. 35, 140-142.Chayen, N.E. (2002) Tackling the bottleneck of protein crystallization in the post-genomic eraTrends in Biotechnol. 20, 98Chayen, N.E. and Saridakis, E. (2002) Protein crystallization for Genomics: Towards HighThroughput Optimisation Techniques. Lead Article Acta Cryst D 58, 921-927.Chayen, N.E. and Helliwell, J.R., (2002) "Protein Crystallization in Microgravity: Are We Reapingthe Full Benefit of Outer Space? Annal. New York Academy of Sciences 594, 591-597.Cianci, M., Rizkallah, P.J., Olczak, A., Raftery, J., Chayen, N.E., Zagalsky, P.F. & Helliwell, J.R. (2002)"The Molecular Basis of the Coloration Mechanism in Lobster Shell: Beta Crustalcyanin at 3.2 AResolution." PNAS 99, 9795-9800.Chayen, NE. and Hilgenfeld, R. (2002) Crystallization Comes of Age Trends in Biotechnol. 20, 320-321.Saridakis, E., Dierks, K., Moreno, A., Dieckmann, M. W. M. and Chayen, N.E. (2002) SeparatingNucleation and Growth in Protein Crystallization Using Dynamic Light Scattering Acta Cryst D 58,1597-1600.Nield, J., Rizkallah, P., Barber, J. and Chayen, N.E. (2003) The 1.45 Å Three-dimensional Structureof C-Phycocyanin from the Thermophilic Cyanobacterium Synechococcus Elongatus J. Struct.Biol. 141, 149-155 (and Front cover).Dunlop, K.V. and Hazes B. (2003) When less is more: a more efficient vapour diffusion protocolActa Cryst. D59, 1797-1800.Saridakis, E. and Chayen, N.E. (2003) Systematic Improvement of Protein Crystals by Determiningthe Supersolubility Curves of Phase Diagrams Biophys J. 84, 1218-1222.Chayen N.E. (2003) Protein Crystallisation for Genomics: Throughput versus Output J. Structuraland Functional Genomics 4, 115-120.D'Arcy, A., MacSweeney, A. and Haber, A. (2004) Practical aspects of using the microbatchmethod in screening conditions for protein crystallization. Methods 34 (3) 323-328.Nneji, G.A. and Chayen, N.E. (2004) A crystallization plate for controlling evaporation in hangingdrops J. Appl. Cryst. 37, 502-503.Chayen, N.E. (2004) Turning Protein Crystallization from an Art into a Science Current Opinion inStructural Biology 14, 577-583.Chayen, N.E. (2005) Methods for separating nucleation and growth in protein crystallizationProgress in Biophysics and Molecular Biology 88, 329-337.Chayen N.E., Saridakis, E. and Sear, R.P. (2006). Experiment and theory for heterogeneousnucleation of protein crystals in a porous medium. Proc. Natl. Acad. Sci. USA. 103: 597-601Chayen, N.E. (2006) “Optimization Techniques for Automation and High throughput” Methods inMolecular Biology, 363, 1, 175-190.Chayen, N.E. “Automation of non-conventional crystallization techniques for screening andoptimization” Chapter 3 in "Macromolecular Crystallography, Conventional and High-throughputMethods" Sanderson, M. and Skelly, J. ed. (Oxford University Press) pp 4-58 2007.Helliwell, J.R. and Chayen, N.E. (2007). “A down to-Earth Approach. Nature 448, 58-59.D’Arcy, A., Villard, F. & Marsh, M. (2007). Acta Cryst. D63, 550–554.Khurshid, S., Govada, L. and Chayen, N.E. (2007) ”Dynamic screening experiments to maximizehits for crystallization” Crystal Growth & Design 7, 2171-2175.Chayen, N.E. and Saridakis, E. (2008) “Protein crystallization: From purified protein to diffractionqualitycrystal” Nature Methods, 5, 147-153.Chayen, N.E. (2009) Rigorous Filtration for Protein Crystallization. J. Appl. Cryst. 42, 743–744.Saridakis, E. and Chayen, N.E. (2009) “Towards a ‘Universal’ Nucleant for Protein Crystallization”Trends in Biotechnology 27, 99-106.Govada, L. and Naomi E. Chayen, N.E. (2009) “Crystallization by Controlled Evaporation Leadingto High Resolution Crystals of the C1 Domain of Cardiac Myosin Binding Protein-C (cMyBP-C)”Crystal Growth & Design 9, 1729-1732.Chayen, N.E. (2009) High Throughput Protein Crystallization. Advances in Protein Chemistry andStructural Biology, 77, 1-22.Bolanos-Garcia, V.M. and Chayen, N. E. (2010) New directions in conventional methods of proteincrystallization. Progress in Biophysics and Molecular Biology 101, 3-12.Khurshid, S., Haire, L.F. and Chayen, N.E. (2010) Automated seeding for the optimization of crystalquality. J Appl. Cryst. 43, 752–756.Boudjemline, et al. (2011) Use of Dual Polarisation Interferometry as a diagnostic tool for proteincrystallisation. Anal. Chem. 83, 7881–7887.Garcia-Caballero et al. (2011). Optimization of Protein Crystallization: The OptiCryst Project. Cryst.Growth Des. 11, 2112–2121.Shaw Stewart et al. (2011) Random Microseeding: A Theoretical and Practical Exploration of SeedStability and Seeding Techniques for Successful Protein Crystallization. Crystal Growth & Design11, 3432–3441.Saridakis et al. (2011) Protein crystallization facilitated by molecularly imprinted polymers. Proc.Natl. Acad. Sci. USA 108, 11081-11086.BooksProtein Crystallisation Bergfors, T.M. ed. 2009 La Jolla, International University Line, USA.“Macromolecular Crystallization and Crystal Perfection” 2010 N.E. Chayen, J.R. Helliwell, andE.H. Snell. Oxford University Press, Oxford, UK.Crystallisation of Biological Macromolecules. 1999 McPherson, A. Cold Spring Harbor LaboratoryPress, Cold Spring Harbor, USA.Crystallization of Nucleic Acids and Proteins. 1999 Ducruix, A. & Giegé, R., eds. Oxford UniversityPress, Oxford.Methods and Results in crystallization of Membrane Proteins. 2003 Iwata, S. ed La Jolla,International University Line, USA.Macromolecular Crystallography Protocols 2006 Doublie, S. ed Humana Press, New Jersey.Macromolecular Crystallography, Conventional and High-throughput Methods 2007 Sanderson,M.R. and Skelly, J. V. ed. Oxford University Press.Protein Crystallization Strategies for Structural Genomics. 2007 Chayen, N. E. ed La Jolla,International University Line, USA.North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com 59

PLATESTRADITIONAL “LINBRO” STYLE PLATES FOR HANGING DROP WITH COVERSLIPS OR SITTING DROP WITH BRIDGES WERE JUST THE BEGINNING.61 24-Well plates61 XRL 24–well plates61 24-well SBS plates61 Grease61 Sitting drop bridges62 Coverslips63 Counter diffusion60 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

24-WELL PLATESXRL 24-WELL PLATESCLASSIC 24-WELL “LINBRO” STYLE PLATEWITH A RAISED LID.PLATESACCESSORIESFOR 24-WELL PLATES.24-Well Plates & AccessoriesXRL PlateThe raised lid, “Linbro style” plate for hanging drop or sitting dropwith bridges.This is the standard plate for manual use and optimization.Sealing GreaseThe Dow Corning standard for all vapour diffusion set-ups.Features of XRL Plate:n Excellent optical properties.n Linbro style footprint.n Raised lid gives ample room for coverslips (22 mm).n Trial pack with CrystalClene Slips available.Features of Sealing Grease:n A non-drying, chemical resistant silicone sealant.n Maintains its consistency from -40°C through 260°C.Sitting Drop BridgesFit both 24-well SBS format plates and ”Linbro style” XRL plates.Features of Sitting Drop Bridges:n Sit neatly and firmly inside the well.n Provide a stable sitting drop platform for vapour diffusion.n Excellent optical properties with polished surfaces for inspection.24-WELL SBS PLATES24-WELL PLATES IN SBS FORMAT.Greiner Combo plateA 24-well SBS format plate which can be used with 18 mm coverslipsfor hanging drop or with bridges for sitting drop.ORDER INFORMATIONDescription Pack CodeXRL Plate 100 MD3-11XRL Plate Trial Pack 40 MD3-11T(includes 1000 22 mm CrystalClene Slips)Greiner 24-well Combo Plate 24 MD3-16Greiner pre-greased 24-well Combo Plate 24 MD3-21ORDER INFORMATIONDescription Pack CodeSealing grease 50g MD6-02Sitting Drop Bridges 100 MD3-17North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com61

PlatesCOVERSLIPSCLENEGLASS COVERSLIPSCOVERSLIPSCRYSTALCLENE SLIPSGERMAN GLASS COVERSLIPS PLAIN ORSILICONISED FOR HANGING DROP EXPERIMENTS.POLYCARBONATE UNBREAKABLE COVERSLIPSDO NOT NEED TO BE SILICONISED FOR HANGINGDROP EXPERIMENTS.Clean, glass coverslips are available in both plain and siliconisedpresentations.Unbreakable, clean polycarbonate coverslips for hanging drop.Whilst plastic coverslips had been used for some time, MolecularDimensions’ introduction of these polycarbonate coverslipssignified a great advance in coverslip technology for hangingdrop crystallization.Features of CleneGlass Coverslips:n Siliconised and plain glass coverslips.n 3 sizes: 18 mm round, 22 mm round and 22 mm square.Features of CrystalClene Slips:n Do not require silanisation.n Unbreakable chemical & solvent resistant plastic coverslips.n Supplied protected in a sandwich film or interleaved.n Guaranteed dust-free.n Guaranteed RNase-free.n Available round and square.ORDER INFORMATIONDescription Pack CodeSiliconised 18 mm round 1000 MD4-02Siliconised 22 mm round 1000 MD4-04Siliconised 22 mm square 1000 MD4-06Plain 18 mm round # 2* 1000 MD4-07Plain 22 mm round # 2 1000 MD4-08Plain 22 mm square # 2 1000 MD4-09Plain 18 mm round # 3* 1000 MD4-18Plain 22 mm round # 3 1000 MD4-19* Thickness of coverslips# 2 = 0.19 – 0.23 mm# 3 = 0.28 – 0.32 mmORDER INFORMATIONDescription Pack Code22 mm square 100 MD4-1022 mm square 1000 MD4-1118 mm round 100 MD4-1218 mm round 1000 MD4-1322 mm round 100 MD4-1422 mm round 1000 MD4-1518 mm square 100 MD4-1618 mm square 1000 MD4-1762Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

COUNTER DIFFUSIONCOUNTER DIFFUSIONCRYSTALHARP THIS IS THE FIRST, AND HENCE UNIQUE DEVICE FORGROWING CRYSTALS IN CAPILLARIES SET IN AN SBSFRAME, ALLOWING TRUE IN-SITU DATA COLLECTION.This capillary plate is based on the successful interface diffusionmethod.Features of Crystalharp:n Apply protein in one simple step.n 48 channels of proven optimal length (30 mm) sampling largeamounts of protein phase space.n No scale up needed – in-situ diffraction – shoot through the plate ormount individual capillaries in standard CryoCaps.n In-situ diffraction means no handling of crystals, no mounting, lessmechanical damage.n Unique capillary material allows data collection at RT and/or flashfrozenin a liquid nitrogen stream (with or without the use of cryoprotectants).n SBS format suitable for high throughput screening.n Simple set up and analysis, seal with everyday crystallography tape.n Easy addition of cryo protectants or derivatives for phasing studies.n Compatible with all storage and imaging systems – no additionalholders needed.n Room for labelling – by hand or with barcodes.PlatesORDER INFORMATIONDescription Pack CodeCrystalHarp Starter Pack* 1 MD11-57-SPCrystalHarp 1 MD11-57CrystalHarp 5 pack 5 MD11-57-5CrystalHarp Capillary Cutting Tool 1 MD7-520CrystalHarp Capillary Tweezers- rubber tipped 1 MD7-521Cryotongs 24 mm 1 MD7-519* The Starter Pack contains 2 CrystalHarp plates, filling and sealing devices and instructionbooklet.References1. Garcia-Ruiz JM, Methods in Enzymology, 1997, Vol. 3682. Cianci M, Helliwell JR, Suzuki A., Acta Crystallogr D, 2008 Dec; 643. Pos KM, Schiefner A, Seeger MA, Diederichs K., FEBS Lett. 2004 Apr 30; 564(3)North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com63

COUNTER DIFFUSIONHIGH QUALITY DIFFRACTION DATA IS A PREREQUISITE TO DETERMINEPHASES OF SUFFICIENT QUALITY AND TO OBTAIN A STRUCTUREFROM A CRYSTALLIZED MACROMOLECULEDr Mareike KurzThe most common method to obtain diffraction datais to take crystals grown by vapour diffusion andmanually manipulate them during the steps of cryoprotection, plunge cooling and mounting for datacollection (1-4) . The manual manipulation steps canbe inappropriate for fragile and difficult to handlecrystals and might even prevent the acquisition ofuseable diffraction data.An alternative is offered by the method of counter diffusion, inwhich the crystallization is performed in long capillaries, usinghighly concentrated precipitating solutions to provokesuccessive nucleation events approaching equilibrium (5-7) . Themethod of counter diffusion, especially in restricted geometry, allowsprotein crystallization under nonconvective conditions and thereforemimics microgravity experiments. During the equilibration of the proteinand precipitating solution a supersaturated spatiotemporal gradientalong the capillary is formed. This creates a continuous nucleation front,which travels along the protein solution and allows crystallization at afavourable concentration at any position in the capillary. This uniquenature of the counter diffusion method has the advantage of a muchbroader screening of variables in one single experiment (Figure 1).Dr Mareike Kurz and Dr Beat Blattmann, University of Zurich, Switzerland.Figure 1. shows the solubility diagram for counter diffusion crystallization. During crystallization, the proteinsolution is driven to the supersaturated region by reducing the solubility of the macromolecule from thesolvent (top). The nucleation moves into the labile region, which reduces the solubility of the protein solutionuntil the metastable region is reached. In a counter diffusion experiment (bottom), the protein and precipitantsolutions are placed in front of each other inside a capillary. The two solutions diffuse against each other,forming a spatial–temporal gradient of supersaturation and a continuous nucleation front that travels alongthe capillary. Figure adapted from (7).64Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

Additionally, the counter diffusion process can be used to screen foroptimal conditions for protein crystal growth, to incorporate stronganomalous scattering atoms as well as to add cryogenic solutions in asingle capillary tube. Moreover, problems related to harvesting crystalsand difficulties in transportation are eliminated.Over the last decades several tools and devices combining counterdiffusion crystallization in capillaries with in-situ diffraction analysis andpost-crystallization treatments have been developed and implemented instructural genomic pipelines. Counter diffusion based crystallization toolsinclude microofluidic devices (8-13) , microcapillary-based microbatchplastic tubings (14, 15) as well as glass capillaries (5-7, 16, 17) . Furthermore,counter diffusion based crystallization devices that couple visualizationand automation for in-situ X-ray diffraction analyses to address highthroughput screening have been developed. Worth mentioning are theCOC based microchannel crystallization plates, which can be assembledto fit in SBS-format frames (18) .ProteinloadingwellsHowever, most methods and tools available to date (for a review see (19) )are not practical, need large amounts of protein and are far from beinguseful for today's state-of-the-art nanolitre scale high throughputcrystallization facilities. The newest capillary counter diffusion basedcrystallization device commercially available is the CrystalHarp (Figure2). In comparison to all other devices, the CrystalHarp (i) allows counterdiffusion crystallization in 48 individual experiments (ii) consumes only500 nl protein solution for one individual capillary experiment, (iii) caneasily be loaded with protein solution and offers simultaneously screeningfor new and optimal growth conditions, incorporation of anomalousscatterers and addition of cryogenic solutions in a single capillary, (iv)allows addition of precipitating buffer using standard dispensing robotsand (v) due to its SBS format can be imaged with standard imagingsystems used for sitting drop vapor diffusion experiments. TheCrystalHarp allows in-situ X-ray diffraction analysis of a crystal by anyconventional X-ray source. Otherwise, an individual capillary can beremoved from the crystallization plate and the crystal can be analyzed360° in situ by X-ray diffraction.An additional advantage of capillary counter diffusion crystallization isthe possibility to perform high pressure freezing, hence omitting thetedious procedure to search for a cryo protectant to avoid hexagonal iceformation. Rapid freezing of aqueous specimens at a pressure of 210 MPa(high pressure freezing) reduces or prevents the formation of cubic orhexagonal ice phases and thus avoids ice rings in X-ray diffractionexperiments (20-23) .Individual precipitation wellsSealing troughsGlass capillaryFigure 2. shows the CrystalHarp plate, designed for 48 counter diffusion crystallization experiments.The indicated protein loading wells, the 48 individual precipitation wells and the four cutting and sealingtroughs to create the single experiments are indicated. Picture taken from (23).References1. Berry, I. M., Dym, O., Esnouf, R. M., Harlos, K., Meged, R., Perrakis, A., Sussman, J. L., Walter, T. S., Wilson, J.,and Messerschmidt, A. (2006) SPINE high-throughput crystallization, crystal imaging and recognitiontechniques: current state, performance analysis, new technologies and future aspects, Acta Crystallogr DBiol Crystallogr 62, 1137-1149.2. Weber, P. C. (1997) Overview of protein crystallization methods, Macromolecular Crystallography, Pt A276, 13-22.3. Pflugrath, J. W. (2004) Macromolecular cryocrystallography--methods for cooling and mounting proteincrystals at cryogenic temperatures, Methods 34, 415-423.4. Garman, E. F., and Owen, R. L. (2006) Cryocooling and radiation damage in macromolecularcrystallography, Acta Crystallogr D Biol Crystallogr 62, 32-47.5. Garcia-Ruiz, J. M., and Ng, J. D. (2007) Counter-Diffusion Capillary Crystallization for High-ThroughputApplications, In Protein Crystallization Strategies for Structural Genomics (Chayen, N. E., Ed.), pp 111–126,La Jolla: International University Line.6. Gavira, J. A., Toh, D., Lopez-Jaramillo, J., Garcia-Ruiz, J. M., and Ng, J. D. (2002) Ab initio crystallographicstructure determination of insulin from protein to electron density without crystal handling, ActaCrystallogr D Biol Crystallogr 58, 1147-1154.7. Ng, J. D., Gavira, J. A., and Garcia-Ruiz, J. M. (2003) Protein crystallization by capillary counterdiffusion forapplied crystallographic structure determination, J Struct Biol 142, 218-231.8. Hansen, C. L., Skordalakes, E., Berger, J. M., and Quake, S. R. (2002) A robust and scalable microfluidicmetering method that allows protein crystal growth by free interface diffusion, Proc Natl Acad Sci U S A99, 16531-16536.9. Zheng, B., Roach, L. S., and Ismagilov, R. F. (2003) Screening of protein crystallization conditions on amicrofluidic chip using nanoliter-size droplets, J Am Chem Soc 125, 11170-11171.10. Zheng, B., Tice, J. D., and Ismagilov, R. F. (2004) Formation of Arrayed Droplets by Soft Lithography andTwo-Phase Fluid Flow, and Application in Protein Crystallization, Adv Mater 16, 1365-1368.11. Zhou, X., Lau, L., Lam, W. W., Au, S. W., and Zheng, B. (2007) Nanoliter dispensing method by degassedpoly(dimethylsiloxane) microchannels and its application in protein crystallization, Anal Chem 79, 4924-4930.12. Gerdts, C. J., Tereshko, V., Yadav, M. K., Dementieva, I., Collart, F., Joachimiak, A., Stevens, R. C., Kuhn, P.,Kossiakoff, A., and Ismagilov, R. F. (2006) Time-controlled microfluidic seeding in nL-volume droplets toseparate nucleation and growth stages of protein crystallization, Angew Chem Int Ed Engl 45, 8156-8160.13. Du, W., Li, L., Nichols, K. P., and Ismagilov, R. F. (2009) SlipChip, Lab Chip 9, 2286-2292.14. Kalinin, Y., and Thorne, R. (2005) Crystal growth in X-ray-transparent plastic tubing: an alternative forhigh-throughput applications, Acta Crystallogr D Biol Crystallogr 61, 1528-1532.15. Yadav, M. K., Gerdts, C. J., Sanishvili, R., Smith, W. W., Roach, L. S., Ismagilov, R. F., Kuhn, P., and Stevens, R.C. (2005) In situ data collection and structure refinement from microcapillary protein crystallization, JAppl Crystallogr 38, 900-905.16. Garcia-Ruiz, J. M. (2003) Counterdiffusion methods for macromolecular crystallization, Methods Enzymol368, 130-154.17. Ng, J. D., Stevens, R. C., and Kuhn, P. (2008) Protein crystallization in restricted geometry: advancing oldideas for modern times in structural proteomics, Methods Mol Biol 426, 363-376.18. Ng, J. D., Clark, P. J., Stevens, R. C., and Kuhn, P. (2008) In situ X-ray analysis of protein crystals in lowbirefringentand X-ray transmissive plastic microchannels, Acta Crystallogr D Biol Crystallogr 64, 189-197.19. Otalora, F., Gavira, J. A., Ng, J. D., and Garcia-Ruiz, J. M. (2009) Counterdiffusion methods applied toprotein crystallization, Prog Biophys Mol Biol 101, 26-37.20. Fourme, R., Girard, E., Kahn, R., Dhaussy, A. C., and Ascone, I. (2009) Advances in high-pressurebiophysics: status and prospects of macromolecular crystallography, Annu Rev Biophys 38, 153-171.21. Kim, C. U., Chen, Y.-F., Tate, M. W., and Gruner, S. M. (2008) Pressure-induced high-density amorphous icein protein crystals, J Appl Crystallogr 41, 1-7.22. Mishima, O., Calvert, L. D., and Whalley, E. (1984) ‘Melting ice’ I at 77 K and 10 kbar: a new method ofmaking amorphous solids, Nature 310, 393 -395.23. Kurz, M., Blattmann, B., Kaech, A., Briand, C., Reardon, P., Ziegler, U., and Gruetter, M. G. (2012) Highthroughputcounter-diffusion capillary crystallization and in situ diffraction using high-pressure freezingin protein crystallography, J Appl Crystallogr 45, 999-1008.North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com 65

HIGH-THROUGHPUTPLATESDESIGNED FOR OPTIMAL DROPPLACEMENT, SEALING, IMAGING ANDCRYSTAL HARVESTINGMODERN4TH GENERATIONPLATES66 High-throughput plates68 MRC Crystallization plates68 Triple drop plates68 MRC Maxi Optimization plates69 Microbatch plates69 In situ diffraction plates72 Hanging drop plates & seals72 Screw top hanging drop plate73 Laminex for LCP73 Sealing sheets and deepwell blocksDr Jan LöweMolecular Dimensions together with Swissci AG are committed toproviding you with the very latest, end-user designed,crystallization plates of consistently high quality. Our unique andongoing relationship allows you to buy with confidence that weprovide the best pricing in the market together with rapid deliveryand customer support you can rely on.66Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

CHOOSING THE RIGHTCRYSTALLIZATION PLATESmall protein drops in relatively large drop wells have atendency to “run” to the sides, especially where there areedges and corners. Round drop positions may therefore bemore favourable than square ones if you are intending to usesmall drops.You will have to observe your drops under a microscope. All trayshave their own characteristic optical properties because they areinjection moulded. Small shapes and corners in plastic trays tend toexhibit dark fields. Higher quality mouldings have better opticalproperties with no flow lines or bubbles.You will wish to recover any crystals that form so the protein dropshould not be too far below the upper surface of the tray, so thatthere is a good shallow angle for approaching the drop with a loopfor cryo mounting.The surface properties of the plastics are also important with respectto drop behaviour. New materials and coatings are being introducedto address such issues as birefringence and hydrophobicity. UVtransparent polymers are essential for viewing fluorescent proteincrystals under UV illumination.Sitting dropThe fourth generation MRC Plate (2 Drop Plate) was designed bycrystallographers at the MRC Laboratory of Molecular Biology inCambridge, UK. The Swissci Triple Drop Plate has similar featureswith 3 protein drop positions. Both are available in polystyrene or UVpolymer. The raised, wide conical wells make crystal retrieval easy,and are excellent for drop placement and viewing. Screening andcrystallogenesis experiments can be performed in one plate (10 nL-5µL drops) with 50-100 µL capacity reservoirs. The plates are easy tonavigate under a microscope with micro-numbered wells. The top ofthe plate has wide surfaces for sealing with tape.EditorialHanging dropIndividual wells of the Screw Top Hanging Drop Plate (below) can beidentified and removed without any disturbance to the growingcrystals inside the plate.The raised wide wells make crystal viewing and mounting especiallyeasy. An allen-key-system allows for smooth removal and subsequentmicroscopic investigation. The plate can be used for screening as wellas harvesting with 10nL-5 µL drop sizes and 50-200 µL reservoirvolumes.OptimizationThe MRC MAXI Plate (left) replaces the need for 24-well sittingdrop plates.Typical volumes are 200-400 µL of reservoir and 1 µL -10 µL drop sizeso you can place larger drops for bigger crystals whilst consumingless screen solution. These plates have all the market leadingfeatures of the MRC plate (Micro-numbering, optically superior wells,optimum sealing area etc.). The SBS format plates are suitable for usewith robots and are available in UV transmissible polymer andstandard polystyrene.MicrobatchThe MRC Under Oil Crystallization Plate was also developed at theMRC Laboratory of Molecular Biology in collaboration with Dr JanLöwe and colleagues.With this plate you can conduct 2 experiments sequentially: first runmicrobatch under oil, then allow the drop to concentrate byevaporation as a second stage with the volatile oil.The new design of the wells allows easy crystal viewing and retrieval.North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com 67

High-Throughput PlatesSITTING DROP PLATESTHE MRCCRYSTALLISATION PLATE THE FOURTH GENERATION CRYSTAL GROWTH PLATE.OPTIMIZATION PLATEMRC MAXI OPTIMIZATIONPLATE – 48 WELLALSO DEVELOPED AT THE MRC (CAMBRIDGE, UK), THENEW MRC MAXI PLATE REPLACES THE NEED FOR 24-WELLSITTING DROP PLATES.Now there is a plate with all the features you have been waitingfor – designed by crystallographers at the MRC Laboratory ofMolecular Biology in Cambridge.Place larger drops for bigger crystals whilst consuming less screensolution.Features of MRC Crystallisation plate:n Raised, wide wells make crystal retrieval easy.n The wells are conical and have a unique lens for exceptional dropplacement and viewing.n Screening and crystallogenesis experiments can be performed in oneplate (10 nL-5 µL drops).n The reservoirs hold volumes from 50-100 µL.n Micro-numbering of each well makes navigating under a microscopeeasy.n The top of the plate has wide surfaces for better sealing with tape.n The plates are designed to the 96-well SBS standard for robotichandling.Molecular Dimensions offers this superior 96-well sitting drop platedesign in two materials; Standard polystyrene, or Optically superior andUV transmissible patented polymer.Features of the MRC Maxi Optimization Plate:n All the market leading features of the standard MRC plate (Micronumbering,optically superior wells, optimum sealing area etc.)n Typical volumes are 200-400 µL of reservoir and 1 µL -10 µL dropsize.n SBS format and available in UV transmissible polymer and standardpolystyrene.ORDER INFORMATIONDescription Pack CodeTrial Pack of 10 MRC Crystallization 1 MD11-00Plates & 10 ClearVue SheetsMRC Crystallisation Plate Polystyrene 100 MD11-00-100MRC Crystallisation Plate Polystyrene 40 MD11-00-40MRC Crystallisation Plate Polystyrene 10 MD11-00-10MRC Crystallisation Plate UV Polymer 100 MD11-00U-100MRC Crystallisation Plate UV Polymer 40 MD11-00U-40MRC Crystallisation Plate UV Polymer 10 MD11-00U-10ClearVue (Sealing) Sheets 100 MD6-01STriple Drop Plate Polystyrene 100 MD11-003-100Triple Drop Plate Polystyrene 40 MD11-003-40Triple Drop Plate Polystyrene 10 MD11-003-10Triple Drop Plate UV Polymer 100 MD11-003U-100Triple Drop Plate UV Polymer 40 MD11-003U-40Triple Drop Plate UV Polymer 10 MD11-003U-10ORDER INFORMATIONDescription Pack CodeMRC Maxi Optimization 10 MD11-004-10Plate PolystyreneMRC Maxi Optimization 40 MD11-004-40Plate PolystyreneMRC Maxi Optimization 100 MD11-004-100Plate PolystyreneMRC Maxi Optimization 10 MD11-004U-10Plate UV PolymerMRC Maxi Optimization 40 MD11-004U-40Plate UV PolymerMRC Maxi Optimization 100 MD11-004U-100Plate UV PolymerMolecular Dimensions is proud to be associated with the manufacturer –Swissci Ag, and only supply their crystallographer approved, high quality, SBS format crystallization plates.68Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

MICROBATCH PLATESMICROBATCHPLATESFOR MICROBATCH UNDER OIL CRYSTALLIZATION.IN-SITU DIFFRACTION PLATESCRYSTALQUICKX-RAY PLATETHE CRYSTALQUICK X PLATE BY GREINERBIOONE -A NEW LOW BIREFRINGENT PLATE SITTING DROP PLATEOPTIMIZED FOR IN-SITU SCREENING.High-Throughput PlatesMRC Under Oil Crystallization PlateDeveloped at the MRC – Laboratory of Molecular Biology incollaboration with Dr Jan Löwe and colleagues.Features of MRC Under Oil Crystallization Plate:n Conduct 2 experiments sequentially.n Run microbatch under oil.n Allow to evaporate as a second stage with volatile oil.n Made from an optically superior polymer.n New design of the wells allows easy crystal viewing and retrieval.n 96-well UV Transparent polymer sbs plate.ORDER INFORMATIONDescription Pack CodeMRC Under Oil Crystallization Plate 100 MD11-41-UVPMRC Under Oil Crystallization Plate 10 MD11-41-UVP-10Terasaki Microbatch plateThe original HLA (Terasaki) 72-well plates for microbatch under oil*.Features of CrystalQuick:n Low-profile plate (8mm thick).n Low Birefringence plastic for improved UV detection.n Collect X-ray images with an 80 degree angular range.n Individual micro-numbering of wells.n SBS format -Compatible with crystallization robots.ORDER INFORMATIONDescription Pack CodeCrystalQuick X-ray Plate 80 MD11-58Crystal Quick X-ray plates (20) 20 MD11-58-20CrystalQuick X-ray Plate Hydrophobic 80MD11-59X-CHIPAn integrated platform for high-throughput microbatch proteincrystallization and in situ X-ray diffraction data collection. TheX-CHIP (X-ray Crystallization High-throughput Integrated Platform)is a novel microchip that has been developed to combine themultiple steps of the crystallographic pipeline from crystallizationto diffraction data collection on a single device to streamline theentire process. The system has been designed for microbatch(under oil) crystal growth screening, inspection, initial X-rayscreening and data collection in a high-throughput fashion. X-raydiffraction data acquisition can be performed directly on-the-chipat room temperature using an in situ approach. The capabilities ofthe chip eliminate the necessity for manual crystal handling andcryoprotection of crystal samples, while allowing data collectionfrom multiple crystals in the same drop. The platform employshydrophilic and hydrophobic concentric ring surfaces on a miniatureplate transparent to visible light and X-rays to create a well definedand stable microbatch crystallization environment.Features of X-Chip:n Grow high-quality well diffracting crystals.n Collect high-resolution diffraction data sets.n Generate interpretable electron-density maps.n Use nanolitre-scale drop volumes.n Can be used directly on in-house systems & synchrotron beamlines.n Offers full automation of the X-ray structure-determination process.ORDER INFORMATIONDescription Pack CodeTerasaki Microbatch Plate 72-well 120 MD3-14Terasaki Microbatch Plate 72-well 270 MD3-14B*For microbatch oils see page 51.ORDER INFORMATIONDescription Pack CodeF6 X-Chip Starter Kit* 8 MD11-70F6 X-Chip Set 8 MD11-71(use with the Caps, Base Plate and Cover from a F6 Starter Kit)F24 X-Chip Starter Kit# 4 MD11-72F24 X-Chip Set 4 MD11-73(use with the Caps, Base Plate and Cover from a F24 Starter KitNorth & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com69

High-Throughput PlatesHIGH THROUGHPUT PLATES & X CHIPSMRC Crystallization PlateMRC Under Oil Crystallization PlateTriple Drop PlateCrystalQuick X-Ray PlateMRC Maxi Optimization PlateTerasaki Microbatch PlateScrew Top Hanging Drop PlateF24 X-Chip & F6 X-Chip70 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

High-Throughput PlatesClearVue Sealing SheetsLaminexOptiClear SealsEasySeal DWBHanging Drop Starter KitSureSeal DWBNorth & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com71

High-Throughput PlatesHANGING DROP PLATES & COVERSHANGING DROP PLATE& UV HANGING DROPSEAL PACKTHIS CONVENIENT PACK OFFERS MASSIVE SAVINGS INBOTH MONEY AND TIME SPENT IN SETTING UP HANGINGDROP CRYSTALLIZATION EXPERIMENTS. THE PACKCONTAINS 100 96-WELL SBS FLAT BOTTOM MICROPLATESAND 100 96-WELL UV.HANGING DROP PLATES & COVERSSCREW TOP HANGINGDROP PLATETHE SCREW TOP HANGING DROP CRYSTALLIZATIONPLATE IS A BREAKTHROUGH FOR PROTEINCRYSTALLIZATION IN A 96-WELL FORMAT. FOR THE FIRSTTIME INDIVIDUAL WELLS CAN BE IDENTIFIED ANDREMOVED WITHOUT ANY DISTURBANCE TO THEGROWING CRYSTALS INSIDE THE PLATE.The contents of this pack represent 9600 hanging drop experimentsfor a third of the price you would pay for the classic 24-well platesand coverslips approach. This pack is equivalent to 400 x 24-wellplates and 9600 coverslips.Hanging Drop Starter Kits:These kits contain all you need to to get started with your hangingdrop crystallization experiments. In these kits you will find 596-well flat bottom plates, 5 sheets suitable for cutting out andretrieving your crystals, and an HT-96 crystallization screen. Youcan choose from any of 6 HT-96 crystallization screens fromclassical approaches to highly efficient screens, screeningcomplexes and membrane proteins.96-Well Flat Bottom Plates:These standard (SBS) flat bottom plates have excellent opticalclarity and are intended for use with UV transparent hanging dropcover sheets.UV Transparent Hanging Drop Cover Sheets:These sheets are contact adhesive coated with clear windows forsetting up hanging drops. They are UV transparent and can easilybe cut to retrieve crystals.ORDER INFORMATIONDescription Pack CodeHanging Drop Plate & UV 100 plates & MD11-60Hanging Drop Seal Pack 100 sheetsHanging Drop Starter Kit 5 plates, 5 sheets MD11-60-30Structure Screen I &II HT-96 + screenHanging Drop Starter Kit 5 plates, 5 sheets MD11-60-33MemStart +MemSys HT-96 + screenHanging Drop Starter Kit 5 plates, 5 sheets MD11-60-36PACT Premier HT-96+ screenHanging Drop Starter Kit 5 plates, 5 sheets MD11-60-40JCSG+ HT-96+ screenHanging Drop Starter Kit 5 plates, 5 sheets MD11-60-41MemGold HT-96+ screenHanging Drop Starter Kit 5 plates, 5 sheets MD11-60-42ProPlex HT-96+ screen96-well Flat Bottom Plates 100 plates MD11-02-10096-well Flat Bottom Plates 10 plates MD11-02-10UV Transparent Hanging 100 sheets MD11-00HDDrop Cover SheetsUV Transparent Hanging 10 sheets MD11-00HD-10Drop Cover SheetsRoller to seal sheets to plates 1 MD11-04Note: Hanging Drop Starter Kits screens are supplied in 96-well Deep Well Blocks with 1 mL of reagents.The plate has been developed together with leadingCrystallographers. It is the result of many years of experience insuccessful robotic high-throughput crystallization.The plate offers unique properties that make it ideal for bothnanolitre crystallization screening and microliter optimization alike.Made from an optically superior polymer (UVP) and with a newdesign of wells, the plate allows easy crystal viewing and retrieval.Features of Screw Top Hanging Drop Plate:n Raised wide wells make the crystal mounting especially easy.n Allen-key-system allows for smooth removal and subsequentmicroscopic investigation / X-ray.n Novel crystal-plug-closure all included in the system.n Easy viewing - The wells are wide and conical and have a lens effectfor perfect illumination.n Micro-numbering ensures you will never loose the location under themicroscope.n The optically superior polymer is UV transmissible and can be used todifferentiate between salt and protein crystals.n Wide partition walls between each well provide a high surface area forvery good sealing.n Integrated advanced quality sealing tape - no need to purchaseadditional parts prior to starting a procedure.n Typical volumes are 50-200 µL of reservoir and 10nl-5 µL drop size.n The complete plate is designed to the 96-well SBS standard for allcommon holders.n The system is fully covered by design and patent protection.ORDER INFORMATIONDescription Pack CodeScrew Top Hanging DropCrystallization Plate 10 MD11-010-HDP72 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

IN LAMINA PLATESLAMINEX FOR LIPIDIC CUBIC AND SPONGE PHASE CRYSTALLIZATION.SEALING SHEETSSEALING SHEETSMOLECULAR DIMENSIONS OFFERS A CHOICE OFOPTICALLY CLEAR SEALING SHEET FOR SEALINGCRYSTALLOGRAPHY PLATES.Since the change in adhesive formulation of CrystalClear Manco ®tape Molecular Dimensions have tested over 20 different tapes fortheir optical clarity, sealing ability and performance in the presenceof aqueous and organic vapours.High-Throughput PlatesLaminexTypically, when used for the crystallization of membrane proteinsin lipidic cubic and sponge phases the phase is dispensed onto asurface and reagents are pipetted onto it allowing the crystallizingagents to diffuse into the lipidic drop.Laminex offers considerable advantages for viewing and imagingsuch crystal growth experiments since the experiment is sandwichedbetween planar surfaces. The optical path creates no aberrationseven when using lipidic cubic phases.The Laminex system is built up from lower support plates of glassor UV transparent plastic with an adhesive spacer*. On to this base,glass, UV transparent plates or films can be applied to create 96chambers for crystal growth.Laminex is a novel and unique plate for growing crystals in lamina.In lamina offers the benefits of capillary spaces without the confinesof introducing reagents and samples into and retrieving crystalsfrom capillaries. By adjusting reagent and sample volumes a varietyof experiments can be performed.If protein and reagent drops are dispensed as discrete, and inseparated positions, vapour diffusion can occur within the spacedefined by the upper and lower plates and the spacer. Similarexperiments can be performed with water soluble proteins in atechnique similar to microbatch if the drops touch or are mixed.ORDER INFORMATIONDescriptionLaminex Glass Base 200 micronLaminex Glass Base 100 micronLaminex Plastic Base 200 micronLaminex Plastic Base 100 micronLaminex Glass CoverLaminex Plastic CoverLaminex Film CoverLaminex Starter Kit 200 micronLaminex Starter Kit 100 micronLaminex Frames†Laminex HT Glass Base 200 micron†Laminex HT Glass Base 100 micronTungsten Carbide ScriberCodeMD11-50MD11-50-100MD11-51MD11-51-100MD11-52MD11-53MD11-54MD11-55MD11-55-100MD11-56MD11-50-HTMD11-50-100-HTMD9-29Swissci LCP KitMD11-005U-20* Laminex Bases are available with either a 200 or 100 micron thickspacer to separate the base from the Cover.† SBS size glass base - does not require a frame.ClearVue Sealing Sheetsn UV transparent.n 135 mm x 77 mm optically clear sealing sheet for 96-wellcrystallization plates.n Full adhesive area with two perforated tabs.n Two end tabs for accurate positioning.n To seal plates, apply gentle pressure after positioning on plate.OptiClear Sealsn Fit MRC Crystallization Plates.n Recommended for “in-drop” dynamic light scattering.n UV transparent.EasySeal Deep Well BlockCheap and chearful way to re-seal Deep Well Blocks.n 130 mm x 80 mm sheet with full adhesive area and back slit to aidbacking removal.n Not for long term storage.SureSeal Deep Well BlockProvides longer lasting seal for crystallization screen storage indeep-well blocks.n 137 mm x 80 mm sheet with 117 mm x 80 mm adhesive area.n Two end tabs.n No residue left behind when removed.Deep Well Blocks (empty)Useful for holding crystallization screen reagents. The same blocksas used in our crystallization screens. Polypropylene 96-well, 2 mLblock, V-bottom block (square well).ORDER INFORMATIONDescription Pack CodeOptiClear Seals for MRC Plates 10 MD6-00-MRCClear Sealing tape (50mm x 66M) 1 roll MD6-01ClearVue Sheets 100 MD6-01SEasySeal DWB 100 MD6-15SureSeal DWB 100 MD6-16Sealing Sheet Applicator 10 MD6-18Empty deep 96-well blocks 50 MD3-18Empty deep 96-well blocks 10 MD3-18-1096 Square Well Storage Block Mat 50 MD3-20North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com73

THE INDUSTRY LEADING FAMILY OFDISPENSERS DESIGNED SPECIFICALLYFOR PROTEIN CRYSTALLOGRAPHYGRYPHON• Versatile and upgradable• 96 Channel Syringe Head• Nano Protein Dispense Head• LCP Syringe Dispense Head• Gryphon LCPFactory appointed and trained salesand service agent for South America.74Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

LCP MIXING STATION• Small and quiet• Quick and easy to load• Automatic settings• Speed adjustments• Easy to cleanPHOENIX• Accurate and fast• Easy to use interface• Non-contact dispensing• Precise volume dispensing• Easy plate selectionNorth & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com75

PROTEIN PRODUCTIONYOUR PROTEIN IS THE MAIN VARIABLE76 Protein expression media80 Animal product free media81 Media optimization kit81 LB Broth82 Turbo Broth82 Superior Broth83 Power Broth83 Hyper Broth84 Glucose M9Y, & nutrient mix84 Augmedium85 LB Booster85 Atholate85 Perk expression rescue kit86 Expressmax86 Labelling media87 SelenoMethionine media87 SILAC media88 Cell culture products88 BRFF-BMZERO89 BRFF-EPM289 BRFF-HPC190 BRFF-P4-8F90 PET Cell dissociation formula91 Freezing media pair91 FNC Coating mix92 HEPES buffered saline92 DMEM / F12 serum freemedium93 IMDM serum free medium93 Bovine pituitary extract94 SFM screening kit94 Protein refolding reagents95 Quickfold refolding kit95 Detergent screening kit96 Cyclodextrin kit96 Dialysis & accessories97 Gebaflex dialysis tubes97 Capillaries97 Paper wicks97 LCP mixing adaptor76 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

MEDIA COMPOSITIONINFLUENCESRECOMBINANTPROTEINACCUMULATIONIN E. COLIDr Sheldon E. BroedelEscherichia coli has been the work-horse of gene expressionfor many years and is the first-line system for producingrecombinant proteins. The reason for this is that manydifferent host-vector systems are readily available, theorganism is simple to culture, it grows rapidly, and recovery of therecombinant protein is relatively straightforward, particularly withthe use of affinity tags. Commercially, E. coli is classified as agenerally recognized as safe organism 1 with a good regulatorycompliance history and has proved to be an economically viablemeans for producing protein products. However, not all proteinsare accumulated to maximal levels in E. coli and productiontypically requires optimization.The goal for optimizing production of recombinant proteins is toproduce the highest amount of functional product per unit volumeper unit time. For E. coli, as any other fermentation system, thelevel of intracellular accumulation of a recombinant protein isdependent on the final cell density and the specific activity of theprotein, or, in other words, the level of accumulation relative tototal protein. Four strategies are typically taken for optimizing theproduction of a recombinant protein. These are: The choice ofculture medium, mode of cultivation, strain improvement, andexpression system control (see Lee 2 , Kleman and Strohl 3 , andGustafsson et al 4 for reviews on optimizing strategies).North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com77

Much of the effort aimed at increasing recombinant proteinproduction in fermentations has been directed at maximizing thebiomass production and often overlooked are the effects of mediacomposition on the expression of specific recombinant proteins.However, it is well known that the production of secondarymetabolites in microbial strains can depend on the composition ofthe medium in which the organism is grown. Despite this, littleattention has been paid to the effects of medium formulation onthe accumulation of recombinant proteins.Statistical methods for developing the best medium formulationfor maximizing the production of metabolites is a well-establishedpractice 5 . In the course of optimizing the production of proteins inE. coli, we developed a set of media formulations that consistentlyincrease the accumulation of recombinant proteins above themore traditional LB Broth recipe. Our observations have been thatnot all proteins are maximally expressed in any one medium.Rather, each protein accumulates to different levels in differentmedia and it has not been possible to predict which mediumwould be most suitable for any given protein. Therefore, wedevised a screen in which the production of a recombinant proteinis examined when the host strain is grown in each of six mediaformulations. This simple and rapid screen allows for the selectionof the most suitable medium for any given protein withoutlaborious screening for the critical factors and subsequentoptimization research.The screen uses six different media to identify the formulationyielding the highest level of target protein production. The mediainclude: Glucose M9Y, LB (Miller) Broth, Hyper Broth , PowerBroth , Superior Broth and Turbo Broth . The first two media aretraditional formulations for cultivating E. coli. The latter fourformulations were developed with the objective of increasing thevolumetric production of recombinant proteins 6 . The host strainharboring the expression vectors is grown in each of the media,expression induced and the accumulation of the target proteinmeasured as a function of total biomass and culture volume.Once the best formulation is identified, the selected medium can beused at whatever fermentation size is needed to achieve the desiredproduction levels. Examples of several proteins screened using thisapproach are shown in Figure 1.The medium composition-dependent accumulation of recombinantproteins has several implications. The observation that eachrecombinant protein is expressed to different levels in different mediasuggests that there is a relationship between the respective proteinand the composition of the medium in which it is produced. In theextreme case, the selection of one medium over another could bethe difference between production and no production. The type ofprotein or its original source does not appear to predict whichcomposition is most suitable. The medium producing the highestlevel of biomass also does not predict product accumulation. In ourlaboratory, Hyper Broth gives the highest biomass yields, yet thismedium does not lead a priori to the highest level of recombinantprotein accumulation. Consequently, a screen of several mediaformulations to determine which yields the best level of productionwould be advisable.Despite the history of employing LB medium for the cultivation ofE. coli, this is not necessarily the medium of choice for the productionof recombinant proteins. LB does not contain a buffering system.Therefore, its use in a fermentation system, particularly with feedbatchor continuous culture protocols, is limited. Typically, bufferedmedia formulations are employed in fermentations and mosttraditional fermentation media are of defined compositions 7,8 . Thus,when LB is used in a bench-scale setting to produce recombinantproteins, a different medium must be employed when the productionis shifted to a fermentor. This requires additional research work todetermine the best medium composition. While it can be argued thatsuch work is necessary when regulatory compliance for commercialproduction is an issue, for most research and development, preclinicalor pre-commercial work, this is an unnecessary expense.Selecting a more suitable medium which is more readily scalable, asis the case with Glucose M9Y, Hyper Broth , Power Broth SuperiorA B C D E F GFigure 1. SDS-PAGE analysis of total protein from each of six different host-vector strains. Each strain was grown in 25 mL of the respective medium to an OD600 of 0.6, expression induced with 1 mM IPTG and a 1 mL sampleremoved at 3 h post-induction. The sample was adjusted to give 1OD/mL and a portion analyzed by SDS-PAGE. Panel A – Strain JM109 without a recombinant protein; Panel B to G – Strains harboring plasmids that yield MalE,GST, GFP, I278, TesA, and LypA proteins respectively. Arrows denote the location of the respective recombinant protein. Marker proteins are shown to the left and right of each set of cellular proteins. From left to right in eachpanel are samples from cells grown in Glucose M9Y, LB (Miller), Hyper Broth, Power Broth, Superior Broth and Turbo Broth.78Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

Broth and Turbo Broth (all of which have a buffering system),reduces the time and effort needed to scale the protein production.The observation that the medium composition can affect theaccumulation of soluble protein is significant. The accumulation ofrecombinant proteins as insoluble products is a major shortcoming tousing E. coli-based expression systems. In the example of LypAproduction, the increased amount of soluble protein, as judged byfunctional activity, did not follow the rule of more total product givesmore soluble product. 8 In fact, the amount of soluble protein couldnot be predicted by the relative level of LypA accumulation. Similarresults have been reported for the Candida albicansglucosyltransferase where Power Broth was found to yield solubleproduct whereas the other media did not. Other approachesreported to increase the relative accumulation of soluble proteininclude lowering the temperature during induction 10,11,12 , reducing theconcentration of inducer 13,14 , using mutant strains which affect proteinfolding 15 , fusing the target protein to a bacterial protein 16,17,18 , andemploying a variety of molecular chaperones 19 . Each of these factorsis believed to affect protein folding in one way or another 20 . It may bethat medium composition may affect protein folding too. Most likelythe effect is through modulation of expression of the chaperones oraccessory proteins involved in protein folding. Since a givenchaperone only works on a subset of proteins it is reasonable tosuggest that for any given recombinant protein the set of chaperoneswhich influences its folding may or may not be at sufficient levels inany given medium. Until all of the chaperone-protein interactionshave been defined, it remains beyond the scope of currentknowledge to predict which set of chaperones are needed tocorrectly fold a protein and thus impossible to determine in advancewhich medium formulation is most suitable for the target protein.Therefore, the best medium for producing a given protein must bedetermined empirically.References1 Guidelines for Research Involving Recombinant DNA Molecules, October 2011,Office of Biotechnology Activities, National Institutes of Health,http://oba.od.nih.gov/oba/rac/Guidelines/NIH_Guidelines.htm.2 Lee, S. Y. 1996. High cell-density culture of Escherichia coli. Tibtech 14:98-105.3 Kleman, G. L. and W. R. Strohl. 1994. Development in high cell density and highproductivity microbial fermentation. Current Opinion in Biotech. 5:180-186.4 Gustafsson, C., Minshull, J., Govindarajan, S., Ness, J, Villalobos, A., and Welch,M. 2012. Engineering genes for predictable protein expression. Prot. Exp. Purif.83:37-46.5 Strobel, R. J. and G. R. Sullivan. 1999. Experimental design for improvement offermentations. In Manual of industrial Microbiology and Biotechnology, A. L.Demain and J. E. Davies, eds., ASM Press, Washington, DC, pp.80-93.6 Broedel, Jr., S. E., Papciak, S. M., and Jones, W. R. 2001. The section ofoptimum media formualtions for improved exrepssion of recombinant proteinsin E. coli. Athena Enzyme Systems Group, Technical Bulletin, Vol. 2, January2001. http://www.athenaes.com/ tech_brief_optimum_media.php.7 Trilli, A. 1986. Scale-up fermentations. In Manual of Industrial Microbiology andBiotechnology, A. L. Demain and N. A. Solomon, eds., ASM Press,Washington, DC, pp. 277-307.8 Hosobuchi, M. and H. Yoshikawa. 1999. Scale up of microbial processes. InManual of Industrial Microbiology and Biotechnology, A. L. Demain and J. E.Daves, eds., ASM Press, Washington, DC, pp.236-239.9 McElheny-Feser, G. and R. L. Cihlar, Georgetown University, personalcommunication.10 Piatak, M., J. A. Lane, W. Laird, M. J. gjorn, A. Wang, and M. Williams. 1988.Expression of soluble and fully functional ricin A chain in Escherichia coli. J.Biol. Chem. 263:4837-4843.11 Schein, C. H., and M. H. M. Noteborn. 1988. Formation of soluble recombinantproteins in Escherichia coli is favored by lower growth temperatures.BioTechnology 6:291-294.12 Cabilly, S. 1989. Growth at sub-optimal temperatures allows the production offunctional antigen-binding Fab fragment in Escherichia coli. Gene 85:553-557.13 Takagi, H. Y. Morinaga, M. Tsuchiya, H. Ikemura, and M. Inouye. 1988. Control offolding of protein secreted by high expression secretion vector, pINIIIompA: 16-fold increase in production of active subtilisin E in Escherichia coli.BioTechnology 6:948-950.14 Takagi, H. Y. Morinaga, M. Tsuchiya, H. Ikemura, and M. Inouye. 1988. Control offolding of protein secreted by high expression secretion vector, pINIIIompA: 16-fold increase in production of active subtilisin E in Escherichia coli.BioTechnology 6:948-950.15 Miroux, B. and J. E. Walker. 1996. Over-production of protein in Escherichia coli:mutant hosts that allow synthesis of some membrane proteins and globularproteins at high levels. J. Mol. Biol. 260:289-298.16 Kapust, R. B. and D. S. Waugh. 1999. Escherichia coli maltose-binding protein isuncommonly effective at promoting the solubility of polypeptides to which it isfused. Protein Sci. 8:1668-1674.17 Forrer, P. and R. Jaussi. 1998. High-level expression of soluble heterologousprotein in the cytoplasm of Escherichia coli by fusion to the bacteriophagelambda head protein D. Gene 224;45-52.18 Sachdev, D. and J. M. Chirgwin. 1998. Order of fusions between bacterial andmammalian protein can determine solubility in Escherichia coli. Biochem.Biophys. Res. Commun. 244:933-937.19 Baneyx, F. 1999. Recombinant protein expression in Escherichia coli. CurrentOpinion in Biotechnology.10:411-421.20 Baneyx. F. 1999. In vivo folding of recombinant proteins in Escherichia coli. InManual of Industrial Microbiology and Biotechnology, A. L. Demain and J. E.Daves, eds., ASM Press, Washington, DC, pp. 551-565.North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com 79

Protein ProductionPROTEIN EXPRESSION MEDIAAthena’s proprietaryExpression Media areworld renowned for theirreliability and superiorperformance in theproduction of recombinantproteins. We also offer thePrime and Prime-olateBroths, unique Animal-Product-Free media.Prime Broths contain soyprotein hydrolysates toreplace casein hydrolysate.The proprietary Atholateconsists of three planthydrolysates to emulatethe amino acid balanceof casein hydrolysate.PROTEIN EXPRESSION MEDIAANIMAL-PRODUCT-FREEMEDIAEACH OF OUR ANIMAL-PRODUCT-FREE MEDIA RECEIVEA SEAL OF APF CERTIFICATION AFTER GOING THROUGHRIGID QUALITY CONTROL TESTING.We are commited to providing the highest-quality researchreagents. For our line of animal-product-free media, this includesinsuring comparable (if not superior) performance to standardmedia, in addition to complying with stringent animal-freemanufacturing requirements. To make it easier for laboratorieslooking for animal-free products in our catalogue, we have placedan indicator next to all products in this catalogue that are eitheranimal-product-free, or that have an APF alternative available.In addition there is an index below of all the APF Certified products.For questions about Animal-Product-Free media or quotes on bulkpackaging, contact us.ORDER INFORMATIONDescriptionAPF Media Optimization Kit (powder)APF Media Optimization Kit (liquid)APF LB Broth (Luria)APF LB Broth (Lennox)APF LB Broth (Miller)Turbo Prime Broth (powder)Turbo Prime Broth (liquid)Turbo Prime-olate (powder)Turbo Prime-olate (liquid)Superior Prime Broth (powder)Superior Prime Broth (liquid)CodeMD12-100-APFMD12-129MD12-131MD12-132MD12-133MD12-110MD12-120MD12-160MD12-170MD12-111MD12-121Superior Prime-olate (powder)MD12-161Superior Prime-olate (liquid)Power Prime Broth (powder)Power Prime Broth (liquid)Power Prime-olate (powder)Power Prime-olate (liquid)Hyper Broth (powder)Hyper Broth (liquid)Glucose M9Y (powder)Glucose M9Y (liquid)Glucose Nutrient MixAtholateMD12-171MD12-112MD12-122MD12-162MD12-172MD12-107MD12-118MD12-108MD12-119MD12-109MD12-13480 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

PROTEIN EXPRESSION MEDIAMEDIAOPTIMIZATION KIT THE MEDIA OPTIMIZATION KIT, ANIMAL-PRODUCT-FREEMEDIA OPTIMIZATION KIT AND NEW PRIME-OLATE APFMEDIA OPTIMIZATION KIT ENABLE RESEARCHERS TODETERMINE THE BEST MEDIA FORMULATION THAT YIELDSTHE HIGHEST AMOUNT OF TARGET PROTEIN.PROTEIN EXPRESSION MEDIALB BROTH & ANIMAL-PRODUCT-FREE LB BROTHWE OFFER THREE VARIATIONS OF THE BASIC NUTRIENT-RICH LB BROTH MEDIA (MILLER, LENNOX, AND LURIA) ASWELL AS THEIR APF CERTIFIED EQUIVALENTS MADE USINGTHE PROPRIETARY BLENDED PLANT PROTEINHYDROLYSATE: ATHOLATE.Protein ProductionSince the production of a given recombinant protein can varysignificantly depending on the type of medium employed, theMOK’s provide four proprietary expression broths, as well asGlucose M9Y, and LB Miller Broth. The APF kit contains APFCertified versions of Athena’s proprietary expression media, andthe Prime-olate Kit contains the new Prime-olate expressionseries, formulated with Athena’s casein hydrolysate replacement,Atholate. Each dry powder kit contains enough mix to prepare 1liter of each of the six media.The ready-to-use liquid kits contain 100 mL of each medium.The formulations each provide an abundance of peptides andpeptones, vitamins and trace elements, and differ only in sodiumchloride content and whether LB Broth & Animal-Product-Free LBBroth they were made with casein hydrolysate or Atholate.The differences allow the researcher to select which formula willprovide the ideal osmotic conditions for their particular bacterialstrain and desired culture conditions.Each LB Broth and APF LB Broth can be custom ordered in bulk.Kit ComponentsStandard Media Optimization KitSuperior Broth, Turbo Broth, Power Broth, Hyper Broth,LB Broth (Miller), Glucose M9Y, Applications ManualAPF Media Optimization KitSuperior Prime Broth, Turbo Prime Broth, Power Prime Broth,Hyper Broth, LB Broth (Miller), Glucose M9Y, Applications ManualPrime-olate Media Optimization KitSuperior Prime-olate, Turbo Prime-olate, Power Prime-olate,Hyper Broth, APF LB Broth (Miller), Glucose M9Y, Applications ManualORDER INFORMATIONDescription Pack CodeMedia Optimization Kit Powder 1 kit MD12-100APF Media Optimization Kit Powder 1 kit MD12-100-APFLiquid Media Optimization Kit 1 kit MD12-128Liquid (ready-to-use)Liquid APF Media Optimization Kit 1 kit MD12-129Liquid (ready-to-use)Prime-olate APF Media Optimization 1 kit MD12-169Kit PowderLiquid Prime-olate APF Media 1 kit MD12-179Optimization Kit Liquid (ready-to-use)FORMULAS (g/L)Ingredient Luria Lennox MillerCasein Hydrolysate / Atholate 15 15 15Yeast Extract 5 5 5NaCl 0.5 5 10ORDER INFORMATIONDescription Pack CodeLB Broth (Luria) Powder 500g MD12-101LB Broth (Lennox) Powder 500g MD12-102LB Broth (Miller) Powder 500g MD12-103LB Broth (Lennox) Liquid (ready-to-use) 5 x 500mL MD12-113LB Broth (Miller) Liquid (ready-to-use) 5 x 500mL MD12-114LB Broth (Miller) Powder (Box of 10) MD12-1301L Single-use-PacketsAPF LB Broth (Luria) Powder 500g MD12-131APF LB Broth (Luria) Powder (Box of 10) MD12-131-S1L Single-use-PacketsAPF LB Broth (Luria) Liquid 5 x 500mL MD12-174(ready-to-use)APF LB Broth (Lennox) Powder 500g MD12-132APF LB Broth (Lennox) Powder (Box of 10) MD12-132-S1L Single-use-PacketsAPF LB Broth (Lennox) Liquid 5 x 500mL MD12-175(ready-to-use)APF LB Broth (Miller) Powder 500g MD12-133APF LB Broth (Miller) Powder (Box of 10) MD12-133-S1L Single-use-PacketsAPF LB Broth (Miller) Liquid 5 x 500mL MD12-173(ready-to-use)North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com81

Protein ProductionPROTEIN EXPRESSION MEDIATURBO BROTH &TURBO PRIME BROTH TURBO BROTH, TURBO PRIME BROTH AND THE NEWTURBO PRIME-OLATE ARE PROPRIETARY MEDIAFORMULATIONS USED FOR THE CULTIVATION OFRECOMBINANT STRAINS OF E. COLI.PROTEIN EXPRESSION MEDIASUPERIOR BROTH &SUPERIOR PRIME BROTH SUPERIOR BROTH, SUPERIOR PRIME BROTH ANDNEW SUPERIOR PRIME-OLATE, ARE PROPRIETARYMEDIA FORMULATIONS FOR IMPROVED EXPRESSIONOF RECOMBINANT PROTEINS IN E. COLI.Both Turbo Prime and Turbo Prime-olate are APF Certified.Turbo Prime-olate is made with Atholate, Athena’s proprietaryblended plant protein hydrolysate. The media improve the yield ofrecombinant proteins and typically produce cell yields 4 to 5 timeshigher than that of LB Broth in shake flask cultures depending on thestrain. They have higher plasmid copy numbers than LB, which canenhance expression. Each medium uses glycerol for a carbon sourceand has a rich nutrient base of amino acids, vitamins, inorganicminerals and trace minerals. The media are buffered at pH 7.2 ± 0.2with potassium phosphate to prevent a change in pH and provide asource of phosphate. These media are ideal as the base forfermentation cultures.The formulations are complex, supplying a nitrogen source, vitamins,and moderate glucose levels. The media are buffered at pH 7.2 ±0.2 to prevent acidification of the medium during metabolism of theglucose. Expression of recombinant proteins should be induced atcell densities twice that of LB Broth. Cell yields are typically about3 to 4 times that of LB Broth. Superior Broth shows the highestplasmid copy numbers among our family of media. This medium is anexcellent choice for large-scale plasmid preparations. Biomassyields will depend on the process and the strain employed.Features of Turbo Broth:n Cell yields up to 5 times higher than that of LB Brothn High plasmid copy numbersn Rich nutrient basen Ideal as fermentation culture baseFeatures of Superior Broth & Superior Prime Broth:n Cell yields up to 4 times higher than that of LB Brothn High plasmid copy numbersn Ideal for large-scale plasmid preparationLB BrothAPF LB BrothTypical Cell Yields for Turbo BrothLB BrothAPF LB BrothTypical Cell Yields for Superior BrothTurbo BrothTurbo Prime Broth0 6 12 18 24 30 (g)Superior BrothSuperior Prime Broth0 6 12 18 24 30 (g)ORDER INFORMATIONDescription Pack CodeTurbo Broth Powder 500g MD12-104Turbo Broth Powder 1L Single-use-Packet (Box of 10)MD12-104-STurbo Prime Broth Powder 500g MD12-110Turbo Prime Broth Powder (Box of 10) MD12-110-S1L Single-use-PacketTurbo Broth Liquid (ready-to-use) 5 x 500mL MD12-115Turbo Prime Broth Liquid (ready-to-use) 5 x 500mL MD12-120Turbo Prime-olate Powder 500g MD12-160Turbo Prime-olate Powder (Box of 10) MD12-160-S1L Single-use-PacketsTurbo Prime-olate Liquid (ready-to-use) 5 x 500mL MD12-170ORDER INFORMATIONDescription Pack CodeSuperior Broth Powder 500g MD12-105Superior Broth Powder1L Single-use-Packet(Box of 10) MD12-105-SSuperior Prime Broth Powder 500g MD12-111Superior Prime Broth Powder1L Single-use-Packet(Box of 10) MD12-111-SSuperior Broth Liquid (ready-to-use) 5 x 500mL MD12-116Superior Prime Broth Liquid(ready-to-use)5 x 500mL MD12-121Superior Prime-olate Powder 500g MD12-161Superior Prime-olate Powder1L Single-use-Packet(Box of 10) MD12-161-SSuperior Prime-olate Liquid(ready-to-use)5 x 500mL MD12-17182 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

PROTEIN EXPRESSION MEDIAPOWER BROTH &POWER PRIME BROTH POWER BROTH, POWER PRIME BROTH, AND POWERPRIME-OLATE ARE PROPRIETARY MEDIUM FORMULATIONSFOR IMPROVED EXPRESSION OF RECOMBINANT PROTEINSIN E. COLI.PROTEIN EXPRESSION MEDIAHYPER BROTH HYPER BROTH IS AN APF CERTIFI ED PROPRIETARYMEDIA FORMULATION FOR IMPROVED EXPRESSION OFRECOMBINANT PROTEINS IN E. COLI. THE MEDIUM IS ACOMPLEX, RICH FORMULATION WHICH SUPPLIES AMINOACIDS, VITAMINS, AND GLUCOSE AT HIGH LEVELS. THISMEDIUM YIELDS THE HIGHEST LEVEL OF BIOMASS OF ALLOUR MEDIA.Protein ProductionThese media are composed of a complex, rich mixture of aminoacids, vitamins, and a carbon source at higher levels than TerrificBroth. The media are buffered at pH 6.8 ± 0.2. Power Prime andPower Prime-olate are APF Certified. For expression ofrecombinant proteins, induction should be done at cell densitiesthree times that of LB Broth. Cell yields are typically 3 to 4 timesthat of LB Broth in shake flask cultures depending on thecharacteristics of the strain. Several reports suggest that PowerBroth and Power Prime Broth can increase the amount of solubleprotein accumulated for otherwise insoluble proteins. This medium isa favorable base for fermentation cultures. Biomass yields willdepend on the process and the strain employed.The medium is buffered at pH 7.3 ± 0.2 and has the highestbuffering capacity of our Expression Media, which permits highglucose levels. For expression of recombinant proteins, inductionshould be done at cell densities three times that of LB Broth. Cellyields are typically 5 times that of LB Broth depending on the strain.Plasmid copy numbers are significantly higher than they are for LBBroth. This medium is suitable as the base for fermentation culturesand is excellent for feed-batch applications. Biomass yields willdepend on the process and the strain employed. Hyper Brothpowder is supplied with a separate Glucose Nutrient Mix.Features of Power Broth & Power Prime Broth:n Cell yields up to 4 times higher than that of LB Brothn Rich nutrient basen Favorable base for Fermentationn CulturesFeatures of Hyper Broth:n Cell yields up to 5 times higher than that of LB Brothn Supplied with Glucose Nutrient Mixn Ideal as fermentation culture baseLB BrothAPF LB BrothTypical Cell Yields for Power BrothLB BrothAPF LB BrothTypical Cell Yields for Hyper BrothPower BrothPower Prime Broth0 6 12 18 24 30 (g)Hyper Broth0 6 12 18 24 30 (g)ORDER INFORMATIONDescription Pack CodePower Broth Powder 500g MD12-106Power Broth Powder(Box of 10) MD12-106-S1L Single-use-PacketPower Prime Broth Powder 500g MD12-112Power Prime Broth Powder(Box of 10) MD12-112-S1L Single-use-PacketPower Broth Liquid (ready-to-use) 5 x 500mL MD12-117Power Prime Broth Liquid5 x 500mL MD12-122(ready-to-use)Power Prime-olate Powder 500g MD12-162Power Prime-olate Powder(Box of 10) MD12-162-S1L Single-use-PacketPower Prime-olate Liquid5 x 500mL MD12-172(ready-to-use)ORDER INFORMATIONDescription Pack CodeHyper Broth Powder 500g MD12-107Hyper Broth Powder1L Single-use-Packet(Box of 10) MD12-107-SHyper Broth Liquid (ready-to-use) 5 x 500mL MD12-118North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com83

Protein ProductionPROTEIN EXPRESSION MEDIAGLUCOSE M9YGLUCOSE M9Y IS AN APF CERTIFIED MINIMAL MEDIAFORMULATION SUPPLEMENTED WITH YEAST EXTRACT FORTHE CULTIVATION OF E. COLI. IT IS BASED ON M9 SALTSWITH A PH OF 6.9 ± 0.2 AND EMPLOYS GLUCOSE AS THECARBON SOURCE.PROTEIN EXPRESSION MEDIAAUGMEDIUM AUGMEDIUM IS A MEDIUM ADDITIVE WHICH CONDITIONSCELLS PRIOR TO INDUCTION DURING A RECOMBINANTPROTEIN EXPRESSION PROTOCOL.)gm/nim/selom µ( ytivitcA c fiicepSBuffering is provided by a sodium-potassium phosphate system.Ammonium chloride provides a nitrogen source. Yeast extract isadded to improve the protein production capacity of the mediumand to permit cultivation of auxotrophic strains by supplying asource of amino acids, vitamins and minerals. Our formulation differsfrom the traditional formulation of Glucose M9, which employscasamino acids. Our experience has shown that yeast extracttypically gives higher yields of recombinant proteins than casaminoacids. Glucose M9Y powder is supplied with a separate GlucoseNutrient Mix.ORDER INFORMATIONDescription Pack CodeGlucose M9Y Powder 500g MD12-108Glucose M9Y Powder1L Single-use-Packet Box of 10 MD12-108-SGlucose M9Y Liquid (ready-to-use) 5 x 500mL MD12-119Augmedium is specifically intended for use with strains in which thetarget protein accumulates as an inclusion body or as an insolubleaggregate. Augmedium is supplied as a powder for preparation of50x concentrated stock solutions.Features of Augmedium:n Medium Additiven Dramatically Increases Level of Expressionn Increases Fraction of Soluble Protein7500Augmedium Dependent Increase in LypA ActivityGLUCOSE NUTRIENT MIX5000GLUCOSE NUTRIENT MIX IS AN APF CERTIFIED MEDIUMSUPPLEMENT USED FOR ENHANCING THE EXPRESSION OFPROTEINS WHEN USING HYPER BROTH OR GLUCOSE M9Y.25000IGP 0x 0.16x 0.31x 0.63x 1.25x 2.5xAugmedium ConcentrationThe powder blend consists of glucose as a carbon source and issupplemented with calcium chloride and magnesium sulphate.Glucose Nutrient Mix comes supplied with both media, but can nowbe purchased separately in individual packets.The graph above shows the level of expression of LypA withincreasing levels of Augmedium. The amount of activity of LypAdirectly correlates with the concentration of Augmedium andshows a significant increase of LypA accumulation. IGP is theisogenic parent strain.ORDER INFORMATIONDescription Pack CodeGlucose Nutrient Mix Powder 150g MD12-109ORDER INFORMATIONDescription Pack CodeAugmedium Powder 100mL Stock MD12-123Augmedium Powder 500mL Stock MD12-12484 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

PROTEIN EXPRESSION MEDIALB BOOSTER LB BOOSTER IS A MEDIUM ADDITIVE THATINCREASES THE LEVEL OF PRODUCTION OFRECOMBINANT PROTEINS. PARTICULARLY SUITABLEFOR IMPROVING THE PRODUCTIVITY OF LB BROTH,LB BOOSTER CAN INCREASE BIOMASS LEVELS BY AFACTOR OF UP TO 5 IN ANY MEDIUM FORMULATION.PROTEIN EXPRESSION MEDIAPERK : PROTEINEXPRESSION RESCUE KITTHE PROTEIN EXPRESSION RESCUE KIT IS INTENDEDTO HELP MAXIMIZE THE AMOUNT OF SOLUBLE PROTEINPRODUCED DURING EXPRESSION OF RECOMBINANTPROTEINS IN E. COLI.Protein ProductionIn addition, the supplement provides essential nutrients selectedfor their ability to increase the relative level of recombinant proteinaccumulation. LB Booster is supplied as a powder and as a readyto-use20x concentrate. LB Booster is a proprietary blend offructose and nutrient supplements.Features of LB Booster:n Increase Biomass Levels by 5 Xn Fructose and Nutrient BlendORDER INFORMATIONDescription Pack CodeLB Booster Powder for 100mL stock 1 packet MD12-125-100LB Booster Powder for 500mL stock 1 packet MD12-126-500LB Booster 20x Liquid (ready-to-use) 500mLMD12-127This comprehensive kit includes samples of our proprietaryExpression Media including Turbo Broth, Superior Broth, PowerBroth, Hyper Broth and two reference media, Glucose M9Y and LB(Miller) Broth. The Expression Media are specially designed toincrease the production of recombinant proteins.The kit also includes the proprietary medium additives: LB*Boosterand Augmedium. LB*Booster is designed to increase the biomassproduction of any expression medium up to 5-fold. Augmedium is amedium pre-conditioner that induces the expression of chaperoneproteins in advance of expression of the target recombinant protein.This can increase the amount of soluble protein accumulated.Features of LB Booster:n Maximize Soluble Proteinn Dramatic Expression Resultsn Includes Media Additive and Preconditioner to Advance ExpressionATHOLATE ATHOLATE IS AN APF CERTIFIED PROPRIETARY BLEND OFPLANT PROTEIN HYDROLYSATES FOR USE IN THEPREPARATION OF MICROBIOLOGICAL MEDIA.Kit Components:Superior BrothTurbo BrothPower BrothHyper BrothLB Broth (Miller)Glucose M9YGlucose Nutrient MixLB BoosterAugmediumApplication ManualAtholate was developed as an Animal-Product-Free alternative tocasein hydrolysate in common expression media such as LB Broth.It is designed to outperform standard casein hydrolysate whilemaintaining the same nutritional characteristics. Our newestexpression broths, the Prime-olate series, feature Atholate as akey component. Atholate is available in bulk quantities.ORDER INFORMATIONDescription Pack CodeAtholate Powder 500g MD12-134Atholate Powder 1kg MD12-134-1Atholate Powder 2.5kg MD12-134-2.5Atholate Powder 10kg MD12-134-10ORDER INFORMATIONDescription Pack CodePERK Protein Expression Rescue 1 kit MD12-135Kit PowderNorth & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com85

Protein ProductionPROTEIN EXPRESSION MEDIAEXPRESSMAX MEDIAAND SCREENING KITATHENA’S EXPRESSMAX SERIES AREFERMENTATION MEDIA FOR THE PRODUCTION OFRECOMBINANT PROTEINS. THE BASAL SALTS AREINORGANIC NUTRIENTS OPTIMIZED TO GIVE THEHIGHEST POSSIBLE GROWTH RATE OF E. COLI ANDPRODUCTION OF RECOMBINANT PROTEINS.The screening kit is intended to aid in formulation developmentby supplying the basal salts along with the carbon (glucose) andnitrogen sources (yeast extract, soy protein hydrolysate andAtholate) which can be combined in different combinations toidentify the composition that gives the highest production ofthe target protein. ExpressMax medium is also available preformulatedfor ease of use. These formulations have beendesigned by Athena’s scientists for maximum production ofrecombinant proteins.Features of ExpressMax Media and Screening Kit:n Fermentation Median Dramatic Expression Resultsn Provides Basal Salts, Media, Nitrogen and Carbon SourcesLABELLING MEDIAReady to use media forincorporation of SeMetinto recombinant proteinsexpressed in E.coli, and arange of media foreukaryotic cell culture forincorporation of Met , Leu,Lys or Arg isotopes orderivatives.Kit Components:ExpressMax Basal SaltsExpressMax Yeast ExtractExpressMax Soy Protein HydrolysateAtholateGlucose Nutrient MixApplication Manual with ExpressMax Forumulations GuideORDER INFORMATIONDescription Pack CodeExpressMax Screening Kit Powder 1 Kit MD12-136ExpressMax Formula 1 Powder 500g MD12-137ExpressMax Formula 2 Powder 500g MD12-138ExpressMax Formula 3 Powder 500g MD12-139ExpressMax Formula 4 Powder 500g MD12-140ExpressMax Formula 5 Powder 500g MD12-141ExpressMax Formula 6 Powder 500g MD12-142ExpressMax Formula 7 Powder 500g MD12-143ExpressMax Formula 8 Powder 500g MD12-144Basal Salts Powder 500g MD12-145Yeast Extract Powder 500g MD12-146Soy Protein Hydrolysate Powder 500g MD12-147Glucose Nutrient Mix Powder 500g MD12-109Atholate Powder 500g MD12-13486 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

LABELLING MEDIASELENOMETHIONINEBASED ON A SYNTHETIC M9 MINIMAL MEDIASUPPLEMENTED WITH GLUCOSE, VITAMINS AND AMINOACIDS WITH THE EXCEPTION OF L-METHIONINE.LABELLING MEDIASILAC MEDIASILAC MEDIA IS PREPARED SPECIFICALLY FOR LABELINGEXPERIMENTS INVOLVING THE USE OF STABLE AMINO ACIDISOTOPES. WE PROVIDE SEVERAL OF THE MOST COMMONLYUSED MEDIA WITHOUT L-ARGININE, L-LYSINE, L-LEUCINEAND L-METHIONINE, THE FOUR MOSTLY COMMONLY USEDHEAVY ISOTOPE LABELED AMINO ACIDS.Protein ProductionThis ready-to-use and time saving product is only available fromMolecular Dimensions.The SILAC medium is sterile filled in 500 ml bottles. Stock solutionsof each of the four non-labeled amino acids are provided with eachbottle of the deficient medium to permit reconstitution of thecomplete medium.Low volume custom production of formulations of other cell culturemedia are also available.Features of SelenoMethionine:n Bacterial growth can be carried out either in the presence ofL-methionine or L-selenomethionine (SeMet).n Plasmid expression constructs should be transformed into amethionine requiring auxotroph.n A glucose-free option is available for use with Novagen’s OvernightExpress Autoinduction System I.How to useDissolve 21.6g of base in 1 litre of DI water and autoclave,Dissolve 5.1g* nutrient mix in 50 mL of DI water and sterile filter andadd to base medium to makeup methionine minus medium.Selenomethionine Solution (250x) - Add 4 mL per litre of medium.Methionine Solution (250x) - Add 4 mL per litre of medium to makeinnoculum medium.*Glucose free versions need only 1.1g.Features of SILAC Media:n Ready-to-use with L-Glutaminen Common Formulationsn Antibiotic-freen Custom Formulations AvailableProduct Specifications:Storage: Store at 4°CStability at 4˚C: 3 - 4 monthspH: 7.3 ± 2Osmolality: 270 - 300mOsMORDER INFORMATIONDescription Pack CodeSelenoMet Medium Complete - 500g + 120g MD12-500(includes wet ice for shipping)2 x 16 mLSelenoMet Medium Complete Glucose 500g + 120g MD12-500-GF- Free (inc. wet ice) 2 x 16 mLSelenoMet base media and nutrients 500g + 120g MD12-501SelnoMet Medium Base 500g MD12-501BSelenoMet Base medium and Nutients 500g MD12-501-GFGlucose FreeSeMet Nutrient Mix 120g MD12-502Glucose free nutrient mix for 120g MD12-502-GFSelenoMet mediaSelonoMethionine solution 250x 16 mL MD12-503(inc. wet ice for shipping)SelenoMethionine Solution 50 mL MD12-503BMethionine solution 250x 16 mL MD12-504(inc. wet ice for shipping)ORDER INFORMATIONDescription Pack CodeDMEM High Glucose 500mL MD12-420DMEM Low Glucose 500mL MD12-421RPMI 1640 500mL MD12-422DMEM/F12 500mL MD12-423MEM 500mL MD12-424Ham's F12 500mL MD12-425McCoy's 5A 500mL MD12-4261,149 mM L-Arg 6 x 0.5mL MD12-416800 mM L-Lys 6 x 0.5ml MD12-417160 mM L-Leu 6 x 2.5ml MD12-418201 mM L-Met 6 x 0.5ml MD12-419References1. Ong, S.-E. and Mann, M. 2006. A practical recipe for stable isotope labeling by amino acids incell culture (SILAC). Nat. Protocols 1(6):2650-2660.North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com87

Protein ProductionCELL CULTURE PRODUCTSThis line of cell cultureproducts includes a seriesof proprietary serum-freetissue culture media inaddition to severalsupplementary productsfor enhancing cell growth,rinsing cells, and cellstorage. All of the serumfreemedia have simplifiedand defined compositionsand can be used topropagate a wide varietyof mammalian cell lines.PROTEIN EXPRESSION MEDIABRFF-BMZERO BRFF-BMZERO IS A COMPLETE SERUMFREEMEDIUM COMPRISED OF ESSENTIAL AMINO ACIDS,VITAMINS, INORGANIC SALTS, TRACE ELEMENTSAND GROWTH FACTORS DESIGNED FOR CULTURINGHUMAN BREAST CELL LINES.BMZERO was developed and has been extensively used forestablishing new epithelial cell lines from explants of humanbreast tissue and to grow immortalized normal breast cell lines.Optimal attached monolayer cell growth can be achieved whenused in conjunction with FNC Coating Mix ® .Features of BRFF-BMZERO:n Designed to grow human breast cellsn Complete serum-free mediumn Antibiotic-freeProduct Specifications:Storage:Store at -80°C, supplied frozenStability at 4˚C: 4 - 6 weekspH: 7.3 ± 2Osmolality: 270 - 300mOsMORDER INFORMATIONDescription Pack CodeBRFF-BMZERO Liquid (ready-to-use) 500mLMD12-40188 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

PROTEIN EXPRESSION MEDIABRFF-EPM2 BRFF-EPM2 IS A COMPLETE SERUM-FREE MEDIUMCOMPRISED OF ESSENTIAL AMINO ACIDS, VITAMINS,INORGANIC SALTS, TRACE ELEMENTS AND GROWTHFACTORS OPTIMIZED FOR CULTURING HUMANOESOPHAGEAL EPITHELIAL CELL LINES.PROTEIN EXPRESSION MEDIABRFF-HPC1 BRFF-HPC1 IS A COMPLETE SERUM-FREE MEDIUMCOMPRISED OF ESSENTIAL AMINO ACIDS, VITAMINS,INORGANIC SALTS, TRACE ELEMENTS, GROWTHFACTORS AND DIHYDROTESTOSTERONE, DESIGNEDFOR ESTABLISHING NEW CELL LINES FROM HUMANPROSTATE TISSUE.Protein ProductionEPM2 has been used for culturing epithelial cells from explantcultures of human skin as well as for culturing a number of humancancer cell lines. Optimal attached monolayer cell growth can beachieved when used in conjunction with FNC Coating Mix ® .HPC1 was developed and has been extensively used forestablishing and maintaining epithelial cell lines from bothbenign prostatic hyperplasia and prostatic carcinoma. Optimalattached monolayer cell growth can be achieved when used inconjunction with FNC Coating Mix ® .Features of BRFF-EPM2:n Optimized for human oesophageal epithelial cellsn Complete serum-free mediumn Antibiotic-freeFeatures of BRFF-HPC1:n Designed to grow human prostate cellsn Complete serum-free mediumn Antibiotic-freeProduct Specifications:Storage:Store at -80°C, supplied frozenStability at 4˚C: 4 - 6 weekspH: 7.3 ± 2Osmolality: 270 - 300mOsMProduct Specifications:Storage:Store at -80°C, supplied frozenStability at 4˚C: 4 - 6 weekspH: 7.3 ± 2Osmolality: 270 - 300mOsMORDER INFORMATIONDescription Pack CodeBRFF-EPM2 Liquid (ready-to-use) 500mL MD12-402ORDER INFORMATIONDescription Pack CodeBRFF-HPC1 Liquid (ready-to-use) 500mL MD12-403North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com89

Protein ProductionPROTEIN EXPRESSION MEDIABRFF-P4-8F BRFF-P4-8F IS A COMPLETE SERUM-FREE MEDIUMCOMPRISED OF ESSENTIAL AMINO ACIDS, VITAMINS,INORGANIC SALTS, TRACE ELEMENTS AND GROWTHFACTORS DESIGNED FOR CULTURING IMMORTALIZEDNORMAL PROSTATIC CELL LINES SUCH AS 267-B1.1PROTEIN EXPRESSION MEDIAPET : CELLDISSOCIATION FORMULAPET IS A SUPERIOR CELL DISSOCIATION REAGENTSPECIALLY FORMULATED FOR USE UNDER “COLD TRYPSIN”CONDITIONS.P4-8F supports the growth of certain established humanprostatic cancer cell lines such as PC-3. Optimal attachedmonolayer cell growth can be achieved when used in conjunctionwith FNC Coating Mix ® .Features of BRFF-P4-8F:n Designed to grow human prostatic cancer cell linesn Complete serum-free mediumThe unique blend of polyvinylpyrrolidone, EGTA, and trypsin in a HBSbase allows the trypsin to retain enzymatic activity under cooltemperatures, unlike standard trypsin which requires a temperatureof 37°C. When used to detach monolayer cultures of epithelial cellsfrom the growth substratum, this reagent yields more viable cellsfrom monolayers grown in serum-free medium than traditionaltrypsin formulas.Features of PET: Cell Dissociation Formula:n Specially-formulated “Cold Trypsin” Reagentn Yields more viable cells than traditional trypsinProduct Specifications:Storage:Store at -80°C, supplied frozenStability at 4˚C: 4 - 6 weekspH: 7.3 ± 2Osmolality: 270 - 300mOsMProduct Specifications:Storage:Store at -80°C, supplied frozenStability at 4˚C: 4 - 6 weekspH: 7.3 ± 2Osmolality: 270 - 300mOsMORDER INFORMATIONDescription Pack CodeBRFF-P4-8F Liquid (ready-to-use) 500mL MD12-404ORDER INFORMATIONDescription Pack CodePET Liquid (ready-to-use) 100mL MD12-40590 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

PROTEIN EXPRESSION MEDIAFREEZINGMEDIA PAIR FREEZING MEDIA PAIR IS A DUAL-CRYOPRESERVATIONSYSTEM USED TO SAFELY STORE SUSPENSIONS OF VIABLECELLS DETACHED FROM MONOLAYER CULTURES USINGTHE “COLD TRYPSIN” METHOD WITH CELL DISSOCIATIONMEDIA, PET.PROTEIN EXPRESSION MEDIAFNC COATING MIX ®FNC COATING MIX ® IS A SPECIALLY FORMULATED SERUM-FREE TISSUE CULTURE REAGENT CONTAININGFIBRONECTIN, COLLAGEN AND ALBUMIN THAT IS USED TOENHANCE THE ATTACHMENT OF ADHERENT CELLS TOPLASTIC FLASKS OR MICROPLATES. THE UNIQUE FORMULACREATES AN EXTRACELLULAR MATRIX THATDRAMATICALLY INCREASES THE RATE OF CELLATTACHMENT TO ANY PLASTIC SUBSTRATUM.Protein ProductionThe pair of solutions thoroughly preserve cells which providessuperior cell viability upon reconstitution. This Freezing Media Pairhas been used extensively to cryopreserve a variety of human celllines grown in serum-free media, including breast and prostatecancer cell lines.Features of Freezing Media Pair:n Dual-cryopreservation systemn Allows for superior cell viabilityThe matrix accelerates monolayer formation, especially whenfastidious cell types such as human prostate and breast epithelialcells are being propagated. Mammalian cell cultures will attach andgrow more effectively on FNC coated plastic surfaces when culturedin the appropriate serum-free medium. We offer several differentserum-free media that when used in conjunction with FNC CoatingMix ® , increase the growth of cell cultures dramatically.Features of FNC Coating Mix ® :n Dramatically Increases Attachment of cellsn Creates Unique Extracellular MatrixProduct Specifications:Storage:Store at -80°C, supplied frozenStability at 4˚C: 4 - 6 weekspH: 7.3 ± 2(for a 1:1 mixture of Solution A and Solution B)Product Specifications:Storage:Store at 4°C, shipped on blue iceStability at 4˚C: 4 - 6 weekspH: 7.3 ± 2Osmolality: 270 - 300mOsMDense monolayer of prostatecancer cells (BRFF-55T) growin HPC1 after application ofFNC Coating Mix to the culture flask.Sporadic cell growth ofprostate cancer cells (BRFF-55T)grow in HPC1 withoutFNC Coating Mix application.The photographs above show the difference in monolayer development with and without FNC Coating Mix ® .BRFF-55T prostate cancer cells were grown in HPC1 medium at the same seed concentration for 3 days at37˚C, 5% CO2. The flask shown on the left was pre-coated with FNC Coating Mix whereas the flask on the rightwas not pre-treated. Through image analysis, there was approximately 85% greater monolayer development inthe flask coated with FNC than in the non-treated flask.ORDER INFORMATIONDescription Pack CodeFreezing Media Pair Liquid 50mL MD12-406(ready-to-use)ORDER INFORMATIONDescription Pack CodeFNC Coating Mix ® Liquid (ready-to-use) 50mL MD12-407North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com91

Protein ProductionPROTEIN EXPRESSION MEDIAHEPES BUFFERED SALINEHEPES BUFFERED SALINE IS AVAILABLE WITH ORWITHOUT PHENOL RED.PROTEIN EXPRESSION MEDIADMEM / F12SERUM-FREE MEDIUMDMEM/F12 IS A SERUM-FREE MEDIUM FORMULATIONFOR GENERAL USE.This is an isotonic saline buffered with Hepes, for use in rinsing cellmonolayers prior to cell dissociation using PET or washingharvested cells.This formulation is a 1:1 blend of DMEM and Ham’s F12 mediasupplied complete and ready to use with L-glutamine, Hepes, BPEand EGF for culturing a wide range of cell types. DMEM/F12 does notcontain phenol red or antibiotics.Features of Hepes Buffered Saline:n Available with or without Phenol Redn Ideal for rinsing cell monolayers prior to cell dissociationProduct Specifications:Storage:Store at -80°C, supplied frozenStability at 4˚C: 4 - 6 weekspH: 7.3 ± 2Osmolality: 270 - 300mOsMProduct Specifications:Storage:Store at 4°C, supplied frozenStability at 4˚C: 12 - 24 monthspH: 7.3 ± 2Osmolality: 270 - 300mOsMORDER INFORMATIONDescription Pack CodeHBS with Phenol Red Liquid 500mL MD12-408(ready-to-use)HBS without Phenol Red Liquid 500mL MD12-409(ready-to-use)ORDER INFORMATIONDescription Pack CodeDMEM / F12 SFM Liquid (ready-to-use) 500mL MD12-41092 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

PROTEIN EXPRESSION MEDIAIMDM SERUM-FREEMEDIUMISCOVE’S MODIFIED DULBECCO’S MEDIUM (IMDM) IS ASTANDARD SERUM-FREE MEDIUM.PROTEIN EXPRESSION MEDIABOVINE PITUITARYEXTRACTBOVINE PITUITARY EXTRACT IS PREPARED FROMBSE-FREE ANIMALS. THE EXTRACTS ARE PREPAREDFROM PITUITARIES HARVESTED FROM ANIMALS INTHE UNITED STATES.Protein ProductionIt is supplemented with bovine pituitary extract and epidermalgrowth factor and contains L-glutamine and Hepes but no phenolred or antibiotics.The pituitaries were obtained only from USDA-inspected facilitieswhere the animals receive pre- and post-mortem inspection andwere found to be free of contagious diseases including foot-andmouthdisease, rinderpest, and contagious bovine pleuropneumonia.There are no known cases of bovine spongiform encephalopathy(BSE) in the herds from which the pituitaries were obtained.Product Specifications:Storage:Store at -80°C, supplied frozenStability at 4˚C: 4 - 6 weekspH: 7.3 ± 2Osmolality: 270 - 300mOsMProduct Specifications:Storage:Store at -80°C, supplied frozenStability at 4˚C: 4 - 6 weekspH: 7.3 ± 2Osmolality: 270 - 300mOsMORDER INFORMATIONDescription Pack CodeIMDM SFM Liquid (ready-to-use) 500mL MD12-411ORDER INFORMATIONDescription Pack CodeBovine Pituitary Extract Liquid 100mg MD12-413Bovine Pituitary Extract Liquid 250mg MD12-414North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com93

Protein ProductionPROTEIN EXPRESSION MEDIASFM SCREENING KIT THE SFM SCREENING KIT IS INTENDED FOR RESEARCHERSSEEKING TO IDENTIFY THE MOST APPROPRIATE SERUM-FREE MEDIA FORMULATION FOR A SPECIFIC CELL TYPE.The kit contains a 100mL sample of each of five serum-free mediaincluding proprietary formulations BRFF-BMZERO, BRFF-EPM2,BRFFP4-8F as well as standard media DMEM/F12, and IMDM.The kit also includes a 25 mL sample of FNC Coating Mix ® foroptimum cell attachment in adherent cell cultures. Each of themedia is complete and ready-to-use.Features of SFM Screening Kit:n Samples of 5 Cell Culture Median Includes FNC Coating Mix ®Kit Components:BRFF-BMZERO DMEM / F12 SFM BRFF-EPM2IMDM SFM BRFF-P4-8F FNC Coating Mix ®BMZERODMEM/F12EPM2PROTEIN REFOLDING REAGENTSWe offer a variety ofprotein refolding reagentsthat include cyclodextrins,detergents, and speciallyformulated refoldingbuffers. QuickFold ProteinRefolding Kit offers asimple and rapid meansof identifying optimalrefolding conditions.It employs a comprehensivematrix design, making iteasy for researchers toscreen a variety of possiblerefolding conditions quickly.P4-8FIMDMThe cell photographs above are of a SFM Screening Kit experiment to determine the optimum serum-freemedia for the production of the prostate cancer cell line 267-B1. The cultures were seeded with 2 x 106cells/mL into each of 5 25cm2 tissue culture flasks pre-coated with FNC Coating Mix ® and containing thefollowing media: BMZERO, EPM2, P4-8F, DMEM / F12 and IMDM. The cells were grown for 24 hours at 37°Cat 5% CO2. The photographs (20x magnification) show the diff erence in cell growth between the diff erentmedia. The cells in the BMZERO media yielded more rapid monolayer development than the other media, withthe least growth in IMDM. These results demonstrate that BMZERO is the optimum medium for the growth of267-B1 cells.ORDER INFORMATIONDescription Pack CodeSFM Screening Kit Liquid 1 kit MD12-415(ready-to-use)94 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

REFOLDING REAGENTSQUICKFOLD PROTEINREFOLDING KIT & BUFFERSTHE QUICKFOLD PROTEIN REFOLDING KIT IS A SCREENINGKIT THAT ENABLES RESEARCHERS TO PINPOINT THECRITICAL FACTORS FOR REFOLDING THEIR PROTEIN IN ASLITTLE AS 1 HOUR.REFOLDING REAGENTSDETERGENT SCREENINGKIT & INDIVIDUALDETERGENTSDETERGENTS HAVE BEEN SHOWN TO AID IN THE PROTEINREFOLDING PROCESS BOTH OF NATIVE AND RECOMBINANTPROTEINS. DETERGENTS ARE USUALLY USED AS A STEP INTHE PROTEIN REFOLDING PROCESS, OFTEN INCONJUNCTION WITH CYCLODEXTRINS.Protein ProductionUnlike many traditional kits, the Quick-Fold Kit employs a fractionalfactorial matrix design that allows the researcher to screen a specificprotein in 15 different buffers quickly and easily. Researchers are ableto examine a wider range of conditions all in a single experiment,simplifying the process of identifying the best buffer composition andmethod for the refolding of a given protein. The QuickFold Kit comeswith enough buffer for 10 refolding experiments, as well assupplemental Dithiothreitol and a Glutathione Redox System. Eachbuffer is available for individual purchase and is supplemented withthe necessary Glutathione Redox System and/or DTT. IndividualBuffers come in 500mL and 1L amounts.Features of QuickFold Protein Refolding Kit & Buffers:n Protein Refolding in as Little as an Hourn Fractional Factorial Matrix Designn Simple, Single-step Screening TechniqueKit Components:Buffers 1 - 15Glutathione, reducedORDER INFORMATIONDTTGlutathione, oxidizedDescription Pack CodeQuickFold Protein Refolding Kit Liquid 1 kit MD1-28The detergent binds to the protein in the dilution stage to form adetergent-protein complex which prevents aggregation of theprotein. A binding agent, such as cyclodextrin, can then be used tostrip away the detergent from the protein, allowing it to properlyrefold. The Detergent Screening Kit provides researchers with 6common detergents (10mL each) for determining the optimumdetergent for a given protein refolding scenario. Individualdetergents can also be purchased in 100mL quantities.Features of Detergent Screening Kit:n Includes 6 Detergents for Comprehensive TestingKit Components:100 mM Triton-X 100 mM POE(10)L100 mM Tween 80 100 mM TTAB100 mM CTAB 100 mM Zwittergent 3-14Individual Refolding Buffers 500 mL 1 LQuickFold Protein Refolding Buffer 1 MD12-612-1 MD12-613-1QuickFold Protein Refolding Buffer 2 MD12-612-2 MD12-613-2QuickFold Protein Refolding Buffer 3 MD12-612-3 MD12-613-3QuickFold Protein Refolding Buffer 4 MD12-612-4 MD12-613-4QuickFold Protein Refolding Buffer 5 MD12-612-5 MD12-613-5QuickFold Protein Refolding Buffer 6 MD12-612-6 MD12-613-6QuickFold Protein Refolding Buffer 7 MD12-612-7 MD12-613-7QuickFold Protein Refolding Buffer 8 MD12-612-8 MD12-613-8QuickFold Protein Refolding Buffer 9 MD12-612-9 MD12-613-9QuickFold Protein Refolding Buffer 10 MD12-612-10 MD12-613-10QuickFold Protein Refolding Buffer 11 MD12-612-11 MD12-613-11QuickFold Protein Refolding Buffer 12 MD12-612-12 MD12-613-12QuickFold Protein Refolding Buffer 13 MD12-612-13 MD12-613-13QuickFold Protein Refolding Buffer 14 MD12-612-14 MD12-613-14QuickFold Protein Refolding Buffer 15 MD12-612-15 MD12-613-15Liquid (ready to use)ORDER INFORMATIONDescription Pack CodeDetergent Screening Kit Liquid 1 kit MD12-601(ready-to-use)100 mM Triton-X Liquid 100 mL MD12-603(ready-to-use)100 mM Tween 80 Liquid 100 mL MD12-604(ready-to-use)100 mM CTAB Liquid 100 mL MD12-605(ready-to-use)100 mM POE(10)L Liquid 100 mL MD12-606(ready-to-use)100 mM TTAB Liquid 100 mL MD12-607(ready-to-use)100 mM Zwittergent 3-14 Liquid 100 mL MD12-608(ready-to-use)North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com95

Protein ProductionREFOLDING AGENTSCYCLODEXTRIN SCREENINGKIT & INDIVIDUALCYCLODEXTRINSCYCLODEXTRINS’ UNIQUE MOLECULAR STRUCTUREGREATLY PROMOTES PROTEIN REFOLDING. CYCLODEXTRINSCAN SUPPRESS PROTEIN AGGREGATION, PROTECT AGAINSTDEGRADATION AND LOSS OF PROTEIN FUNCTION.They also bind well to detergents (aggregation suppressors), whichallows for easy separation of the detergent from the protein,encouraging proper refolding. Different cyclodextrins act andrespond differently to different protein structures. Because of this,we provide 3 of the most common cyclodextrins in a simple screeningkit so that the ideal conditions for refolding a given protein can befound quickly and easily. The Cyclodextrin Screening Kit can beparticularly useful when paired with the Detergent Screening Kit.Individual cyclodextrins are also available for purchase.Features of Cyclodextrin Screening Kit& Individual Cyclodextrins:n Includes 3 Cyclodextrins for Comprehensive TestingKit Components:100 mM β-cyclodextrin50 mM α-cyclodextrin100 mM methyl-β-cyclodextrinORDER INFORMATIONDescription Pack CodeCyclodextrin Screening Kit Liquid 1 kit MD12-602(ready-to-use)100 mM β-cyclodextrin Liquid 100 mL MD12-609(ready-to-use)DIALYSIS & OTHER ACCESSORIESConvenient, low volumetubes for dialysis andelectroelution. Capillariesfor room temperaturemounting. Wicks forremoving excess reagents.An adaptor for fitting twosyringes together formixing lipidic cubic phases.100 mM methyl-β-cyclodextrin Liquid 100 mL MD12-610(ready-to-use)50 mM α-cyclodextrin Liquid 100 mL MD12-611(ready-to-use)96 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

DIALYSISGEBAFLEXDIALYSIS TUBESGEBAFLEX TUBES PERFORM CONVENTIONAL DIALYSISIN ONE TUBE FROM AS LITTLE AS 10 µL.ACCESSORIESOTHERACCESSORIESA SELECTION OF GLASS CAPILLARIES, PAPER WICKS ANDLCP MIXING ADAPTOR SETS.Protein ProductionFeatures of GeBAflex Tubes:n 1KDa to 50KDa molecular weight cut off (MWCO).n Can be used with both proteins and nucleic acids.n High recovery (typically 70% from electro-elution).n Ideal for:– conventional dialysis of small volumes of sample.– extraction of proteins from native gels for assaying.– recovery of protein and removal ofampholytes from samples extractedfrom iso-electric focussing gels.– MALDI-MS sample preparation(30% cheaper than otherpreparation methods).Borosilicate Glass CapillariesA selection of capillary sizes for room temperature mounting ofcrystals.Paper Wicks (sterile)A variety of wicks for wicking away excess reagents fromcrystallization experiments and removing mother liquor fromcapillaries.ORDER INFORMATIONDescription Volume MW Cut-Off Pack Size CodeGebaFlex Tube-Mini10- 250 µL 6-8K 10 MD6-18-1010- 250 µL 6-8K 30 MD6-18-3010- 250 µL 12-14 K 10 MD6-19-1010- 250 µL 12-14 K 30 MD6-19-3010- 250 µL 25 K 5 MD6-20-5Lipidic-Cubic Phase Mixing Adapter SetAn adapter set for mixing your own Lipidic cubic phases.Includes, screw-ended adapting tube and an adapter for fitting twoGas-tight 250/250 µL (and 250/25 µL) Hamilton syringes together(NOT INCLUDED).GebaFlex Tube-Midi50 -800 µL 3500 10 MD6-21-1050 -800 µL 3500 30 MD6-21-3050 -800 µL 6-8K 10 MD6-22-1050 -800 µL 6-8K 30 MD6-22-3050 -800 µL 1 K 5 MD6-23-5GebaFlex Tube-Maxi0.1 – 3 mL 3500 15 MD6-24-150.1 – 3 mL 6-8 K 15 MD6-25-150.1 – 3 mL 12-14 K 15 MD6-26-150.1 – 3 mL 25 K 5 MD6-27-50.1 – 3 mL 50 K 5 MD6-28-5ORDER INFORMATIONDescriptionGlass Capillary Tubes Borosilicate 0.1 mmGlass Capillary Tubes Borosilicate 0.2 mmGlass Capillary Tubes Borosilicate 0.3 mmGlass Capillary Tubes Borosilicate 0.4 mmAssorted wicks (130)Coarse wicks (30)Medium wicks (50)Fine wicks (50)Lipidic-Cubic Phase Mixing Adapter SetCodeMD4-20MD4-21MD4-22MD4-23MD6-03MD6-04MD6-05MD6-06MD6-17North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com97

CRYOCRYSTALLOGRAPHYTHE INTRODUCTION OF STANDARDS FOR SAMPLE MOUNTING ANDROBOTIC SAMPLE CHANGERS HAS FUELLED BOTH HIGHERTHROUGHPUT AND HIGH QUALITY DATA COLLECTION102 LithoLoops102 Elliptical LithoLoops102 Mesh LithoLoops103 CryoMount Sets104 Magnetic CryoVials &CryoCaps105 DataMouse Pro105 Cryoprotectants109 CryoProtX110 Dewars110 Cryogenic shippers,& storage dewars112 EMBL/ESRF samplechanger kits114 CryoTools & accessories98 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

RADIATION DAMAGEIN MACROMOLECULARCRYSTALLOGRAPHYIntensitydecreaseLoss of diffractionProf Elspeth F. GarmanLoss of measuredreflection intensities atroom temperature dueto radiation damage.Incomplete data from crystalsDepartment of Biochemistry,University of Oxford, UKThe advent of highly intense wiggler and undulator beamlinesfed from synchrotron sources has reintroduced the age-oldproblem of X-ray radiation damage in macromolecularcrystallography (MX) even for crystals held at cryogenictemperatures (100 K). Unfortunately, such damage to macromolecularcrystalline samples during the experiment is a problem that is inherentin using ionizing radiation to obtain diffraction patterns and haspresented a challenge to MX since the beginning of the field. Forroom-temperature (RT) data collections, it often necessitates the useof many crystals to assemble a complete data set, because thecrystalline order of the sample is damaged and decreases during theexperiment and thus the diffracted intensity fades. The root cause ofthis damage is the energy lost by the beam in the crystal owing toeither the total absorption or the inelastic scattering of a proportion ofthe X-rays as they pass through the crystal. The measure of this energyloss is the ‘dose’ measured per mass of the sample, given in SI units ofgray (Gy; 1 Gy = 1 J/kg). In MX, dose measurements are generally ofthe order of a million grays (1 MGy or 100 Mrad) and the programmeRADDOSE can conveniently be used to compute the absorbed dosefor a protein crystal (Murray et al. 2004, Paithankar et al. 2009).Up until the 1990s, MX data were almost exclusively collected at RT,where the recommended practice was to monitor the intensity I0 of astrong reflection as the experiment proceeded and to discard thecrystal once the intensity had dropped to 0.85I0, or at the very worst0.70I0 if the particular crystals were in very short supply (Blundell andJohnson 1976).Now most data are collected at 100 K: starting in the 1990scryocooling techniques for MX blossomed and were made technicallymore accessible for routine use in MX because of two pivotaldevelopments: the loop-mounting method (Teng 1990), in which theprotein crystal is held by surface tension in a film of liquid ‘cryo-buffer’across a small-diameter (1 mm down to 0.1 mm) nylon, fibre or plasticloop, and the availability of reliable open-flow unpressurized cryostats(Cosier & Glazer 1986) with flexible stainless-steel hosing to supply aNorth & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com99

stream of cooled gaseous nitrogen at a stable temperature ofaround 100 K with which to surround the sample during datacollection. Initially, problems with the technique included iceformation within and outside the crystal and an increase in mosaicspread, particularly when cryocooling protocols were notoptimized. Methods for improving the data quality obtainablewere soon developed (Rodgers 1997; Garman & Schneider 1997;Garman 1999) and there was widespread adoption of thetechnique. In fact it has been estimated that over 90% of allprotein structures are now determined at cryo-temperatures.However, recent developments at some synchrotrons which allowcrystallization plates to be mounted on beamlines (Axford et al.2012), and experiments which have shown that RT damage can beoutrun using the new very fast detectors (Owen et al. 2012), haveprompted renewed interest in RT collection.The advantages of cryocooling for MX are a very significantreduction in the rate of radiation damage; the use of a mountingtechnique (the loop) that is usually more gentle than the capillarymethod historically used for RT collection; the fact that higherresolution data can more easily be obtained because the crystalorder is preserved for longer; a lower background in the diffractionexperiment as it is not necessary to enclose the crystal in a glass,quartz or plastic tube to prevent dehydration; that fewer crystals(and thus a lower quantity of protein) are required for a project;that crystals can be shipped ahead of time to the synchrotron(more or less) safely; and that crystals can be flash-cooled when inpeak condition for future use before they start to degrade in thecrystallization drop.These positive aspects of cryocooling commonly outweigh thedisadvantages. The latter include the requirement for expensivecryostat cooling equipment, a frequent increase in crystal mosaicspread (but not necessarily if the cryoprotection concentration andcrystal handling are carefully optimized), the need to invest timefor optimization of cryo-buffers and cooling protocols, and the factthat there are as yet no protocols that guarantee success, althoughprogress is being made in this direction.The improvement in dose tolerance for a crystal held at 100 Kcompared with a crystal irradiated at RT has been estimated to beapproximately a factor of 70 on average (Nave & Garman, 2005).A cryoprotected crystal prior to X-rayexposure and crystals after exposure.The band shows the path of the X-raybeam and shows solvated electronsproduced by the X-ray beam.Thus, cryocooling is clearly a highly effective mitigation strategy.However, radiation damage is now routinely observed at synchrotronsin cryocooled crystals and the experimenter would be wise to beaware of the artifacts that can be produced. An experimental doselimit of 30 MGy for the total intensity to drop to 0.70I0 has beenobtained (Owen et al. 2006). Crystals might not survive until this limit,but in the absence of electron scavengers, are not expected toexceed it. Our current knowledge on radiation damage in MX, asreflected in the published literature, is summarized in Garman (2010),Ravelli & Garman (2006), and an article entitled ‘A beginner’s guide toradiation damage’ Holton (2009).Systematic studies of this phenomenon have identified two separateindicators of damage as a function of dose: global and specificdamage. The former manifests as a loss of the measured reflectionintensities (particularly at high resolution), expansion of the unit-cellvolume, increasing values of the measure of the internal consistencyof the data which quantifies the difference between reflectionintensities that should ideally be the same (Rmeas), an increase in boththe scaling B factors for the data and the atomic B values of therefined structure, rotation of the molecule within the unit cell andoften (but not always) an increase in mosaicity. Visible differences inthe samples as the experiment proceeds, including colour changes,are also observed. On warming of the sample following irradiation,bubbles of gas, now proposed to be hydrogen (Meents et al. 2010)and perhaps some CO2, are emitted and discolouration of the sampleis common.Of more direct relevance to the biological interpretation of structuresthan the global indicators mentioned above is the fact that specificstructural damage to particular covalent bonds is observed to occurin a reproducible order in many proteins (Weik et al. 2000; Burmeister2000; Ravelli & McSweeney 2000): first disulfide bridges elongate andthen break, then glutamates and aspartates are decarboxylated,tyrosine residues lose their hydroxyl group and subsequently thecarbon–sulfur bonds in methionines are cleaved. Covalent bonds toheavier atoms such as C—Br, C—I and S—Hg are also ruptured.Clearly, it is not feasible to monitor the specific structural damageduring the experiment, since the refined structures are required.However, it is known that this damage often occurs well before thereis any obvious degradation of the diffraction pattern. Metal centresare also reduced very swiftly by the X-ray beam and increasingly thiscan be monitored using an on-line microspectrophotometer(McGeehan et al 2009) during the X-ray experiment.Radiation damage in MX is an increasingly important and limitingproblem for several reasons. Firstly, as the diffraction experimentproceeds, creeping non-isomorphism occurs on three simultaneousfronts: the unit-cell volume increases, there is often movement of theprotein molecule within the unit cell, and structural changes areinduced by the damage, so that the protein conformation is changingduring the measurements. This non-isomorphism is thought to be amajor cause of unsuccessful MAD (multiple-wavelength anomalousdispersion) structure determinations, since by the time the second orthird wavelength is collected, the cell and atomic structure can havechanged such that the reflection intensities are significantly altered.This effect can obscure the anomalous signal required for structuresolution. It has been calculated that a 0.5% change in all threedimensions of a unit cell would change the intensity of a 3 Åreflection by 15% (Crick & Magdoff, 1956) so the MAD/SAD phasingsignals (typically 4-8% signal) would be completely overwhelmed bysuch a volume increase.100Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

Secondly, the radiation-sensitivity of some crystals at 100 K meansthat it is not possible to collect a complete data set from a singlecrystal and data must be merged from several (or many) of them tomeasure all the unique reflection intensities. Although this wasroutinely the case when data were collected at RT, mostcrystallographers have become accustomed to being able tomeasure all unique reflections from just one cryocooled crystal. Useof multiple crystals to assemble a complete data set in generalincreases the errors arising from non-isomorphism, therebypotentially reducing the ease of structure solution as well asincreasing the mounting/ dismounting time burden.Finally, the radiation-damage-induced structural changes can affectthe apparent biological properties of the macromolecule under study.Enzyme mechanisms can involve redox of susceptible residues, sospecial care is required when interpreting structures that may havebeen modified by X-ray damage during the data collection. Forinstance, irradiation can change the oxidation state of metal ions instructural/ active sites from that in their native state and cause thedecarboxylation of glutamate and aspartate residues. X-ray-inducedstructural changes can also be misleading in studies of intermediates.In such circumstances, separating radiation damage from anenzymatic mechanism can be extremely difficult and can cast doubton the validity of biological conclusions drawn from crystal structures(Ravelli & Garman, 2006).In summary, radiation damage ultimately results in lower resolutionstructures, failed MAD structure solutions and sometimes theinaccurate interpretation of biological results if no controlexperiments are carried out to account for radiation-damage artifacts.It is thus an issue to be taken seriously by the structural biologist.HEWL Asp52 damage. Difference map Fo-Fc4.7 MGy 13.8 MGyDisulphide HEWL Cys6-27, Fo-FcReferencesAxford D, Owen RL, Aishima J, Foadi J, Morgan AW, Robinson JI, Nettleship JE,Owens RJ, Moraes I, Fry EE, Grimes JM, Harlos K, Kotecha A, Ren J, Sutton G,Walter TS, Stuart DI & Evans G (2012) In situ macromolecular crystallographyusing microbeams. Acta Cryst. D68, 592–600.Blundell T, Johnson LN (1976) Protein Crystallography, Academic Press, New YorkBurmeister WP (2000) Structural changes in a cryo-cooled protein crystal owing toradiation damage. Acta Cryst. D56: 328–341Cosier J, Glazer, AM (1986) A Nitrogen-Gas-Stream Cryostat for General X-rayDiffraction Studies. J. Appl. Cryst. 19: 105–107Crick FHC, Magdoff BS (1956) The theory of the method of isomorphousreplacement for protein crystals.I. Acta Cryst. 9: 901–908Garman EF, & Schneider TR (1997) Macromolecular cryocrystallography. J. Appl.Cryst. 30: 211–237Garman E (1999) Cool data: quantity AND quality. Acta Cryst. D55: 1641–1653Garman EF (2010) Radiation damage in macromolecular crystallography: what is itand why should we care? Acta Cryst. D66: 339–351Holton JM (2009) A beginner's guide to radiation damage. J. Synchrotron Rad.16: 133–142Meents A, Gutmann S, Wagner A, Schulze-Briese C (2010) Origin andtemperature dependence of radiation damage in biological samples at cryogenictemperatures. Proc. Natl Acad. Sci. USA, 107: 1094–1099NcGeehan, J, Ravelli RBG, Murray JW, Owen RL, Cipriani F, McSweeney S, WeikM, Garman EF (2009) Colouring Cryo-Cooled Crystals: on-linemicrospectrophotometry. J. Synchrotron Rad.) 16, 163-172.Murray JW, Garman EF, Ravelli RBG (2004) X-ray absorption by macromolecularcrystals: the effects of wavelength and crystal composition on absorbed dose. J.Appl. Cryst. 37: 513–522Nave C, Garman EF (2005) Towards an understanding of radiation damage incryocooled macromolecular crystals. J. Synchrotron Rad. 12: 257–260Owen RL, Rudino-Pinera E, Garman EF (2006) Experimental determination of theradiation dose limit for cryocooled protein crystals. Proc. Natl. Acad. Sci. USA,103: 4912–4917Owen RL, Axford D, Nettleship JE, Owens RJ, Robinson JI, Morgan AW, Dore´AS, Lebon G, Tate CG, Fry EE,Ren J, Stuart DI & Evans G (2012) Outrunning free radicals in room temperaturemacromolecular crystallography. Acta Cryst. D68, 810–818.Paithankar KS, Owen RL, Garman EF (2009) Absorbed dose calculations formacromolecular crystals: Improvements to RADDOSE. J. Synchrotron Rad. 16:152–162Ravelli RBG, McSweeney S (2000) The 'fingerprint' that X-rays can leave onstructures. Structure, 8: 315–328Ravelli RBG, Garman E (2006) Radiation damage in macromolecularcryocrystallography. Curr. Opin. Struct. Biol. 16: 24–629Rodgers DW (1997) Practical Cryocrystallography. Methods Enzymol. 276:183–203Teng T-Y (1990) Mounting of crystals for macromolecular crystallography in afreestanding thin-film. J. Appl. Cryst. 23: 387–391Weik M, Ravelli RBG, Kryger G, McSweeney S, Raves ML, Harel M, Gros, P.,Silman I, Kroon J, Sussman JL (2000) Specific chemical and structural damageto proteins produced by synchrotron radiation. Proc. Natl Acad. Sci. USA, 97:623–6284.7 MGy 13.8 MGyNorth & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com101

Cryo CrystallographyLITHOLOOPSLITHOLOOPS MACHINE-MADE LOOPS OF PRECISE AND REPRODUCIBLEDIMENSIONS HERALDED A REAL BREAKTHROUGH INCRYOCRYSTALLOGRAPHY AND HAVE RAPIDLY TAKEN OVERAS THE NEW STANDARD. PROTEIN WAVE CORP. (JAPAN)DEVELOPED A PROCESS BASED ON LITHOGRAPHY ANDEXCLUSIVELY LICENSED LITHOLOOPS TO MOLECULARDIMENSIONS.LITHOLOOPSELLIPTICALLITHOLOOPSFOR MOUNTING NEEDLE-SHAPED CRYSTALS.Features of LithoLoops:n Improved data collection statistics – Etched from polyimide resinyielding practically zero background diffraction.n Save money and time – re-usable with precise and reproduciblediameters allowing easy matching of loop size to crystal, one pack allthe correct size.n Easier Fishing – Fast and simple retrieval of crystals from drop, evenin viscous solutions. Low hydrophobicity minimises volume of cryoprotectantretained.n Best of both worlds – very strong yet flexible in the drop, rigidityincreases upon freezing giving solid mechanical stability during datacollection.n Choice of size and shape – choose round, elliptical (for needles) ormesh (down to 20 micron) for fragile or micro-crystals.Mounted LithoLoops offer excellent reproducibility with standarddistances from the base of the loop to the end of the mounting pin,making crystal centering easier. LithoLoops are mounted on astandard 18 mm stainless steel pin. The pin has an outside diameterof 0.635 mm to fit all available Cap and Top-Hat mounting systems.ORDER INFORMATIONDescription Diameter CodeMounted LithoLoops 0.02 mm MD7-130Mounted LithoLoops 0.04 mm MD7-131Mounted LithoLoops 0.06 mm MD7-132Mounted LithoLoops 0.08 mm MD7-133Mounted LithoLoops 0.1 mm MD7-134Mounted LithoLoops 0.15 mm MD7-135Mounted LithoLoops 0.2 mm MD7-136Mounted LithoLoops 0.25 mm MD7-137Mounted LithoLoops 0.3 mm MD7-138Mounted LithoLoops 0.4 mm MD7-140Mounted LithoLoops 0.5 mm MD7-142Mounted LithoLoops 0.6 mm MD7-143Mounted LithoLoops 0.7 mm MD7-144Mounted LithoLoops 0.8 mm MD7-145Mounted LithoLoops 1.0 mm MD7-147Mounted LithoLoop Sampler Pack (NEW)Mounted LithoLoop Sampler Pack-CUSTOMMD7-212MD7-212-CUSTOM* Sampler pack contains 2 each of the following 0.06mm, 0.2mm, 0.4mm Round LithoLoops, 0.02 x 0.2mmElliptical LithoLoops and 0.2mm 20 micron Mesh LithoLoops.Elliptical LithoLoops:For mounting needle-shaped crystals.ORDER INFORMATIONDescription Diameter CodeMounted Elliptical LithoLoopsMounted Elliptical LithoLoopsMounted Elliptical LithoLoopsMounted Elliptical LithoLoopMESHLITHOLOOPSPERFECT FOR VERY SMALL AND/OR FRAGILE CRYSTALS,OFFERING MORE SUPPORT.Ideal for "fishing" microcrystals out of drops and for supportingflat "plate-like" crystals. In addition, if several microcrystals arepresent in the drop they will often separate into individual spacesin the Mesh LithoLoop, ideally positioned for alignment on amicro-focussed beamline.ORDER INFORMATIONDescription Size CodeMounted Mesh LithoLoops 0.1 mm - 20 micron0.02 x 0.1 mm MD7-1500.02 x 0.2 mm MD7-1510.02 x 0.3 mm MD7-1520.05 x 0.5 mm MD7-153Mounted Elliptical LithoLoop 0.1 x 0.5 mm MD7-154Mounted Mesh LithoLoops 0.2 mm - 20 micronMounted Mesh LithoLoops 0.2 mm - 40 micronMD7-192MD7-194MD7-195Mounted Mesh LithoLoops 0.2 mm - 60 x 80 micron MD7-196Mounted Mesh LithoLoops 0.3 mm - 20 micronMounted Mesh LithoLoops 0.4 mm - 40 micronMD7-198MD7-201102Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

LITHOLOOPSCRYOMOUNTSETSSETS OF MOUNTED LITHOLOOPS MOUNTED IN CRYOCAPSAND MAGNETIC CRYOVIALS.Cryo CrystallographyEach CryoCap is DataMatrix etched and etched with the size of the LithoLoop.These sets are provided in a handy storage box.ORDER INFORMATIONDescription Diameter CodeCryoMount LithoLoop Set 0.02 mm MD7-130CMCryoMount LithoLoop Set 0.04 mm MD7-131CMCryoMount LithoLoop Set 0.06 mm MD7-132CMCryoMount LithoLoop Set 0.08 mm MD7-133CMCryoMount LithoLoop Set 0.1 mm MD7-134CMCryoMount LithoLoop Set 0.2 mm MD7-136CMCryoMount LithoLoop Set 0.3 mm MD7-138CMCryoMount LithoLoop Set 0.4 mm MD7-140CMCryoMount LithoLoop Set 0.5 mm MD7-142CMCryoMount LithoLoop Set 0.6 mm MD7-143CMCryoMount LithoLoop Set 0.7 mm MD7-144CMCryoMount LithoLoop Set 0.8 mm MD7-145CMCryoMount LithoLoop Set 1.0 mm MD7-147CMCryoMount Elliptical LithoLoops 0.02 x 0.1 mm MD7-150CMCryoMount Elliptical LithoLoops 0.02 x 0.2 mm MD7-151CMCryoMount Elliptical LithoLoops 0.02 x 0.3 mm MD7-152CMCryoMount Elliptical LithoLoop 0.05 x 0.5mm MD7-153CMCryoMount Set Elliptical LithoLoops 0.1 x 0.5 mm MD7-154CMCryoMount LithoLoop Set Mesh 0.1 mm MD7-192CMCryoMount Set Mesh LithoLoop 0.2 mm 20 micron MD7-194CMCryoMount Mesh LithoLoops 0.2 mm - 40 micron MD7-195CMCryoMount Mesh LithoLoops 0.2 mm - 60 x 80 micro MD7-196CMCryoMount Set Mounted Mesh LithoLoops 0.3 mm - 20 micronCryoMount Set Mounted Mesh LithoLoops 0.4 mm x 40 micronCryoMount LithoLoop Set Sampler PackCustom LithoLoop CryoMount SetMD7-198CMMD7-201CMMD7-215CMMD7-CUSTOMNorth & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com103

Cryo CrystallographyCRYOCAPS AND VIALSMAGNETIC CRYOVIALSAND CRYOCAPSTHE EU FUNDED PROGRAMME SPINE (STRUCTURAL PROTEOMICS IN EUROPE),(EMBL-GRENOBLE, F.CIPRIANI, R.RAVELLI, F.FELISAZ) HAS BEEN OF GREATSERVICE TO THE PROTEIN CRYSTALLOGRAPHY COMMUNITY.They defined a terminology and specification for the components of the samplemounting system introducing the standards necessary for robotic sample manipulation.Mounted LithoLoops, CryoCaps and Magnetic Vials, strictly follow SPINE specifications.In particular, tighter engineering tolerances improve precision and reliability for robotichandling and stability for use with micro-crystals.Features of CryoCap:n Rounded edges avoid potential blocking during transfers.n Made from ferromagnetic stainless steel (420).n Minimum quantity of material to reduce the freezing/melting/dryingcycle when the sample holder is transferred.n The pin hole diameter and depth are designed to hold 18 mm pinsprecisely and securely.n Available both plain or with a human readable code and a DATAMATRIX code for automated sample recording.n Codes are laser engraved so that they do not rub off.n Compatible with existing goniometer mounts.n When used with 18 mm pins the sample holder length (cap + pin +loop) is 22 mm.Features of Magnetic CryoVials:n Rounded edges avoid potential blocking during transfers.n A ferromagnetic disc at the bottom of the vials for "positive" vialmanipulation with a magnet.n Magnetic ring polarity, holds the CryoCap, locates in a EMBL/ESRFSample Changer Basket and recovers the CryoCap from thegoniometer mount.n Vent hole to avoid nitrogen pressure build up.n Manufactured to tight engineering tolerances for reliable robotichandling.ORDER INFORMATIONDescription Pack CodeCryocaps 50 MD7-400CryoCaps with Data Matrix 50 MD7-401Magnetic CryoVial 50 MD7-402Magnetic CryoVial with CryoCaps 10 MD7-403Magnetic CryoVial with CryoCaps 50 MD7-404Magnetic CryoVial with CryoCaps 10 MD7-405with data matrixMagnetic CryoVial with CryoCaps 50 MD7-406with data matrixCryoPins (18mm) 50 MD7-410Superglue for CryoPins 1 MD7-410-GLUENotched metal tubes (50) 10 MD7-412104Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

CRYOCAPS AND VIALSTHEDATAMOUSE PROTHIS COMPACT, HIGH-SPEED READER CAN READ ANYDATA-MATRIX CODE.CRYOWARECRYOPROTECTANTSCRYSTALS, EITHER DRY OR ALONG WITH THEIR MOTHERLIQUOR, CAN BE PLACED ON A MICROSCOPE SLIDE ANDCOATED WITH THE CRYO-PROTECTANT.Cryo CrystallographySpecifically, it can be used to read the Data-Matrix code onCryoCaps. The hand held reader is supplied with a softwareinstallation CD. The reader has less than 1 second read time.Features of The DataMouse Pro:n Small foot print.n One simple USB connection required for both power and data.n Data can be sent to any application and can be linked to aspreadsheet or database.n The CryoCap ID is instantly displayed on your computer screen foridentification, or for sample data entry.n Compatible with Hampton & MAR caps.n Can read caps under liquid nitrogen (thin layer of liquid nitrogen).N.B. not compatible with Apple Macs.Excess cryoprotectant can be removed by wicking. Once placed inthe cryostream the cryoprotectant becomes glass-like and protectsthe crystal.Available in 100 mL or 250 mL quantities (where specified).Small molecular weight PEGS (ideal as cryoprotectants) can befound on p37.ORDER INFORMATIONDescription Concentration CodeGlycerol 100 % MD2-100(250)-65Ethylene glycol 100 % MD2-100(250)-60Paraffin Oil 100 %/ 100 mL MD2-03Trehalose 40 % w/v MD2-100(250)-109Xylitol 70 % w/v MD2-100(250)-1102-methyl-2,4-pentanediol 100 % MD2-100(250)-24(MPD)See our CryoProtX screen on page 109.NOZZLE ALIGNMENT TOOLThe Nozzle Alignment Tool has been designed and utilised by Dr EGarman and other practising cryocrystallographers for the exactcentering of a crystal sample in the nitrogen gas stream of anOxford CryoSystems Cooler. Accurate alignment of the stream isvital to avoid ice formation, since it ensures that the crystal staysin the coldest part of the stream as it rotates. Designed for usewith Cryostream Coolers with a serial number greater than 075.ORDER INFORMATIONDescription Pack CodeThe DataMouse Pro 1 MD7-13with Vial RackORDER INFORMATIONDescriptionNozzle alignment toolCodeMD7-07North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com105

THE INDUSTRY LEADING FAMILY OFCOOLERS DESIGNED SPECIFICALLYFOR PROTEIN CRYSTALLOGRAPHYCOBRA• Superior nozzle design• Broad temperature range• Fast cooldown time• Superior stability• Control, monitoringand data loggingFactory appointed and trained salesand service agent for the Americas.106Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

CRYOSTREAM• Fast cool-down• Low LN2 consumption• Variable flow• Highly accurate mapping• Elegant designDESKTOP COOLER• No liquid nitrogen required• Compact and self-contained• Base temperature of 170 K• Very low maintenance• Cryopad softwareNorth & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com107

OVERVIEW OF THE SYSTEMATICAPPROACH TO CRYSTALLIZATIONAND DATA COLLECTIONDr Enrico A. Stura and Laura VeraCEA, iBiTec-S, Service d’Ingénierie Moléculaire desProtéines (SIMOPRO), Gif-sur-Yvette, F-91191, France.PEG concentration & effect on crystallizationOnce you have decided to pursue screening various PEGs at different pHcan be tested 2 to accelerate the screening process and to understandthe trends as a function of pH and PEG molecular weight. While solubilityscreening with PEG is normally instantaneous, salts will requireequilibration for at least 8 hours.Another analysis that can be done quickly is a salt plus PEG screen. In thisway information about certain effects of salt can be obtainedinstantaneously. About 15 minutes after mixing a PEG screen, (such as thePEG conditions from the Stura Footprint Screen #1) you will have dropsthat have precipitate and drops that do not. Now carry out the samescreen adding extra salt (500 mM NaCl or 150-200 mM Li2SO4) to eachreservoir solution before mixing the protein and reservoir solutions. Ifthere is too little salt the protein will tend to precipitate, as salt is added itwill remain soluble (salting in) and as further salt is added it will precipitate(salting out). As a result you will get more or less precipitation. If addingextra salt has a solubilizing effect, a third test can be carried out with evenmore salt (1-4 M NaCl or 1-6 M Li formate). (Avoid high concentrations ofLi2SO4 with high molecular weight PEGs as it will lead to phase separation).Too much precipitation is not good and a drop that does not becomesupersaturated is also useless. This information will help you select whichscreens to set up to maximize your chances of getting crystals.Several steps are required to go from a proteinsample to a crystal structure. The first step is todetermine if the protein is sufficiently concentrated.You can test this easily by taking 1 µl of proteinsolution, adding 1 µl 20% polyethylene glycol (PEG)4,000, 0.2 M imidazole malate, pH 7.0 (solution2C from the Stura Footprint Screen 1 #1, MD1-20)without mixing. If precipitation is observed under amicroscope within 15 minutes, the sample isprobably still suitable for a full scale screen, even ifthe protein is at a concentration as low as 2 mg/ml.Importance of salt in crystallizationThe amount of salt in the protein solution is a very important parameterto consider in vapour diffusion experiments. The reservoir solution shouldhave an equal or higher amount of salt compared to the protein solutionto ensure that drops mixed with an equal volume of protein and reservoirsolutions will shrink during equilibration. During screening any drop thatappears too big compared to the others after equilibration, should beexamined carefully to understand why. You should regularly check yourprotein stock solution for precipitate. If a precipitate is observed, firstcheck for the presence of crystals hidden within the precipitate, then addsalt and change the pH to improve solubility using the knowledgeacquired from the PEG/salt screen to move to more appropriate salt andbuffer conditions that improve solubility. After having done this screeningwill need to be repeated.108Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

Other points to considerYou should also remember to make sure that the buffer the protein isin is compatible with the screening you intend to carry out, and anyglycerol in the protein solution will also need to be matched in thereservoir. 3ScreeningOnce any type of crystalline material has been obtained, screeningcan be carried out together with seeding. You should streak seed 4 allpreviously set up drops. Set up drops with reservoir solutions close tothose that have given the crystalline precipitate and streak seedimmediately the PEG conditions. Repeat the seeding the next day.Screening more finely should lead to better crystals similar to thosealready obtained.To find something new, a different screen can be carried out. Youshould seed all drops immediately for PEG conditions and the nextday for salt conditions. You may obtain an epitaxial jump 5 that maylead to new polymorphs. 6 If you already have a structure, but want toimprove the resolution this may be possible 7 by re-formulating theprecipitants and using additives in the reservoir to potentially alterthe packing.The importance of good cryoprotectionAnother reason for poor diffraction could be a loss in crystallineorder during the cryoprotection step. A good cryoprotectantsolution stabilizes the crystalline state and prevents ice formationduring flash-cooling. A good definition of an appropriatecryoprotectant is where crystals do not dissolve or crack when left inthe cryoprotectant solution for five minutes.A cryoprotectant composition cannot exactly match thecrystallization precipitant unless you have crystallized in a cryosaltor a cryosolution. A simple way to mix a cryosolution was beenpresented at ICCBM-14 8 (CryoProtX MD1-61). It consists of multicomponentmixtures that neither increase nor decrease proteinsolubility but, provide extended stabilization during cryoprotection toallow longer soak periods without crystal cracking or dissolving. Thegeneral practice of using a single anti-icing chemical, such asglycerol, a molecule able to enhance protein solubility, often leads tocrystals dissolving. Using MPD instead can lead to crystals cracking,but by mixing various components together a balance is achieved.CRYOPROTXCRYOPROTX BETTER CRYOPROTECTION FOR QUALITY DIFFRACTION.FINDING A CRYOPROTECTANT THAT GIVES THE OPTIMUMIN TERMS OF DIFFRACTION LIMIT, MOSAICITY ANDCRYSTAL LONGEVITY CAN BE A LABORIOUS ANDFRUSTRATING TASK.The traditional binary approach of soaking in differing amounts ofsingle compounds consumes both potentially good crystals andoften results in making do with the first “ok” result.Ideally in a balanced cryoprotectant the mild solubility enhancingeffect of the glycols should be counteracted by the addition of MPDor DMSO that can act as precipitants. Therefore, an approach usinga complex mixture of glycols and cryoprecipitants should yieldimproved results.CryoProtX achieves this by allowing the user to create a high qualitycryoprotectant from the nine provided intelligent CryoMixes,reducing both the time and crystal consumption whilst increasingthe likelihood of successful cryocooling of the crystal for the highestquality data.Features of CryoProtX:n Preserve the highest diffraction potential of crystals with lowermosaicities.n CryoMixes are provided ready-to-use for single step cryosoaking.n Ideal for PEG and low salt crystallization hits.n Includes pre-made buffers for pH screening.n Vastly reduce the chances of crystal cracking or dissolving.n Provided with additives that may improve crystal longevity atsynchrotron radiation sources.n Fully customizable for individual projects.n Ideal for heavy atom, ligand or back soaks.n Suitable for screening with low affinity, low solubility inhibitors.Developed and formulated in thelaboratory of Dr. Enrico Stura,CryoProtX is manufactured anddistributed under an exclusivelicence with CEA Life SciencesDivision, Saclay, France.Cryo ProtectantsReferences1. Stura E.A., Nemerow G.R., Wilson I.A (1992) Strategies in the crystallization of glycoproteins and proteincomplexes. J. Cryst. Growth 122:273–285 http://dx.doi.org/10.1016/0022-0248(92)90256-I.2. Stura E.A. (1999) Strategy 3: Reverse Screening. In “Crystallization of Proteins: Techniques, Strategies andTips. A laboratory manual” (Bergfors T. ed.) International University Line, pp. 113–124.3. Vera, L., Czarny, B., Georgiadis, D., Dive, V., Stura, E.A. (2011) Practical Use of Glycerol in ProteinCrystallization. Cryst. Growth & Des. 11:2755–2762 http://dx.doi.org/10.1021/cg101364m.4. Stura E.A., Wilson I.A. (1991) Applications of the streak seeding technique in protein crystallization. J.Cryst. Growth 110:270–282 http://dx.doi.org/10.1016/0022-0248(91)90896-D.5. Stura, E.A., Charbonnier, J.-B. Taussig, M.J. (1999) Epitaxial jumps. J. Cryst. Growth 196:250-260http://dx.doi.org/10.1016/S0022-0248(98)00832-X.6. Vera, L., Antoni, C., Devel, L., Czarny, B., Cassar-Lajeunesse, E., Rossello, A., Dive, A. & Stura, E. A. (2013).Screening using polymorphs for the crystallization of protein-ligand complexes.Cryst. Growth & Des.(ICCBM-14, Special issue; Manuscript ID: cg-2012-01537n).7. Ménétrey, J., Perderiset, M., Cicolari, J., Houdusse, A. & Stura, E.A. (2007) Improving Diffraction from3 to 2 Å for a Complex between a Small GTPase and Its Effector by Analysis of Crystal Contacts andUse of Reverse Screening. Cryst. Growth & Des. 7:2140–2146 http://dx.doi.org/10.1021/cg700698d.8. Vera, L., Stura, E.A. (2013)Strategies for protein cryocrystallography. Cryst. Growth & Des.http://dx.doi.org/10.1021/cg301531f.ORDER INFORMATIONDescription Pack CodeCryoProtX46 x 1.5 mL kit MD1-61CryoProtX- Cacodylate-Free46 x 1.5 mL kit MD1-61-CFCryoProtX Single Reagent 1.5 mL 1.5 mL MDSR-61-North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com 109

Cryo CrystallographyDEWARSDEWARSMETAL OR FOAM HOLDERS FOR LIQUID NITROGEN.SHIPPERSCRYOGENIC SHIPPERS,REFRIGERATORS ANDLONG-TERM STORAGEDEWARSFOR STORAGE OR SHIPPING OF PROTEIN CRYSTALSOR CRYOGENIC SAMPLES.Foam DewarsThese new dewars offer improved safety, durability, economy andperformance.Cryogenic DewarsCX (CryoExpress)- Environmental and Biological Shippers.The CX100 CryoExpress Dry Shipper is designed to safely transportsamples at cryogenic temperatures. The CX100 CryoExpress can also besupplied with a shipping case.XT Series (Extended Time) - Refrigerators designed for long-termstorage of materials at cryogenic temperatures.HC Series (High Capacity) - Refrigerators for storing large quantitiesof materials at cryogenic materials.LD Series (Liquid Dewars) Dewars for storing and dispensing smallamounts of liquid nitrogen.ORDER INFORMATIONDescriptionFoam Dewar Value Pack Large (1 std, 1 tall, 1 large)Standard Foam DewarSmall Foam DewarTall Foam DewarLarge Foam DewarFoam Dewar Value Pack (1 small, 1 std, 1 large)CodeMD7-34MD7-35MD7-36MD7-37MD7-38MD7-39600 mL Dewar Full Base Low Form Full Aluminium MD7-8001000 mL Dewar Full Base Low Form MD7-801Full Aluminium hemispherical1000 mL Dewar Full Base w/Metal Handle MD7-802850 mL Dewar low form full base cylindrical MD7-804ORDER INFORMATIONDescriptionLiquid Nitrogen Tank HT SeriesCX100 Dry shipperData Logger for CX100 Dry shipperCX100 dewar lidCX100 Shipping CaseCodeMD7-20MD7-21MD7-21-DATALOGGERMD7-21-LIDMD7-22110 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

Cryo CrystallographyORDER INFORMATIONDescriptionCodeCryo Express (CS Series) Shippers & AccessoriesCX100 Dry shipper (4 L)MD7-21CX100 Shipping CaseMD7-22CXR100 Cryogenic Shipperwith Replaceable absorbantMD7-43CXR100 Replaceable absorbent setMD7-43-9C30CXR500 Dry Shipper (7 L)MD7-46CXR500 Shipping CaseMD7-47Poly Carb 100 Box for CXR500 Dry ShipperMD7-46-BOXES5 shelf rack for CXR500 Dry Shipper MD7-46-RACKData logger for CXR500 Dry shippersMD7-49Data Logger for CX100 Dry shipperMD7-21-DATALOGGERExtended Time (XT) Cryogenic dewars & accessoriesXT-10 Extended Time Cryogenic dewar (10 L) MD7-41XT-20 Extended Time Cryogenic dewar (20 L) MD7-42XTL3 Extended Time Cryogenic dewar (3 L)MD7-48Roller Base for XT-20 (XT) Cryogenic dewar MD7-42-RBLow Liquid Alarm for XT20MD7-72Canister for XT-20 dewarMD7-518Canister for XT-10 dewarMD7-518ORDER INFORMATIONDescriptionHigh Capacity (HC) dewarsHC20 (HC) Dewar (20 L)HC34 (HC) Liquid Nitrogen Storage Dewar (34 L)HC35 (HC) Liquid Nitrogen Storage Dewar (35 L)VHC35 (HC) Liquid Nitrogen Storage Dewar (35 L)HC34/HC35 ESRF Canister Storage CanisterHC35 ESRF Basket Storage SystemLow Level Alarm for HC34 dewarLow Level Alarm for HC35 dewarHC34 roller baseHC 35 roller baseVHC35 roller baseLiquid Dewars (LD Series)LD4 cryogenic dewar (4L)LD10 cryogenic dewar (10L)Classic 25 (25 L)Roller Base for Classic 25Laboratory Systems (LS Series)LS750 35L Liq N2 storage system for racks(750 samples)Roller for LS Series 35L Liq N2 storagesystem for racksCodeMD7-73MD7-450MD7-44MD7-45MD7-44-511MD7-44-512MD7-74MD7-74MD7-450-ROLLERMD7-44-ROLLERMD7-45-ROLLERMD7-71MD7-75MD7-76MD7-76-ROLLERMD7-750-LSMD7-750-ROLLERNorth & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com111

Cryo CrystallographyAUTOMATIC SAMPLE CHANGERTHE EMBL/ESRFAUTOMATIC SAMPLECHANGER KITSDESIGNED FOR LOCAL STORAGE AND TRANSPORTATION OFCRYO MOUNTED SAMPLES AND TRANSFER OF SAMPLESTO THE EMBL/ESRF ROBOTIC SAMPLE CHANGER. FOR USEAT ESRF, SC3 AND IRELEC CATS SAMPLE CHANGERS.These kits provide the baskets to hold the individual samplestogether with all tools and accessories needed to successfully shipsamples for remote data collection.Sample Changer Starter KitContents:n 5 Baskets - Holds 10 SPINE* CryoCaps in Magnetic CryoVials and isuniquely identified with a DataMatrix and Human readable code.n 1 Canister – to fit in CX100 Dry Shipper - Cat# MD7-21 for transport– holds 5 Baskets.n 1 Plug - when placed above the top Basket in the Canister preventsmovement of the Baskets in the CX100 during transit. This works onlyif the canister is full (5 Baskets).n 1 Transfer Support - for loading and unloading vials from Basketsunder liquid nitrogen.n 1 pair of Vial Tongs, 90 degrees to transfer vials to and from theBaskets.n 1 pair of Basket Tongs to transfer Baskets in and out of the Canister.ESRF/EMBL Sample Changer Complete KitSample Changer Starter Complete KitContents, as above, including the following:n 1 pair of CryoTongs for SPINE Caps - to transfer cryocooledcrystals to and from the nitrogen cold stream.n 1 pair of 45 degree Vial Tongs for SPINE Caps - for Cryomanipulation of cryocooled crystals.n 1 Cryo Wand - for use with SPINE magnetic caps.* SPINE standard is 18 mm pin length (22 mm sample holder length).Sample Changer BasketSample Changer Transfer SupportORDER INFORMATIONDescriptionESRF Sample Changer STARTER KitESRF Sample Changer STARTER Kit with CryoCaps & VialsESRF Sample Changer COMPLETE KitESRF Sample Changer COMPLETE Kit& CryoCaps and CryoVialsESRF Sample Changer BasketESRF Sample Changer CanisterESRF Sample Changer Transfer SupportESRF Sample Changer Canister PlugESRF Sample Changer Vial Tongs, 90 degESRF Sample Changer Basket tongsCodeMD7-500MD7-501MD7-502MD7-503MD7-510MD7-511MD7-512MD7-513MD7-514MD7-515ORDER INFORMATIONDescription Pack CodeCryoTongs for SPINE Caps 1 MD7-12Magnetic Cryo Wand for SPINE Caps 1 MD7-411with Suregrip handleVial Tongs, 45 Degrees 1 MD7-516Cryotool Set 1 MD7-517Magnetic 45 degree bent angle 1 MD7-522Cryo Wand for SPINE CapsCryoTongs 24 mm 1 MD7-519112 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

Sample Changer CanisterCryo CrystallographySample Changer Basket Tongs45° Vial Tongs90° Vial TongsCryo WandCryo TongsNorth & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com113

Cryo CrystallographyCRYOTOOLSCRYOTOOLS &CRYO ACCESSORIESTOOLS FOR MANIPULATING CRYOCAPS AND CRYOVIALS.Vial Tongs 45°:For transfer of vials to and from cryocanes and generalcryomanipulation of cryocooled crystals.Cryo Wand:Magnetic cryowand with magnetic plunger for use with SPINEmagnetic caps. Now with SureGrip handle.Cryocanes aluminiumCryo Tongs:Suitable for use with 18 mm (22 mm sample holder length) SPINEstandard caps and pins to transfer cryocooled crystals to and from thenitrogen cold stream. Precision machined and hand-finished StainlessSteel Cryo Tongs manufactured to CE standards. Use these Cryotongswith 45° Vial tongs to manipulate magnetic CryoCaps and Vials.CryoTool Set (includes one each of the above):Cryoware marker setA complete set of Cryotools for manipulating SPINE caps (CryoCaps)and Magnetic CryoVials presented in their own handy carry case;includes CryoWand, CryoTongs and 45° Vial Tongs.Cryogenic vial holderWhite polycarbonate cryogenic vial holder. Allows single-handed vialmanipulation. Bottom of each well interlocks with base of vial. Takes 50vials in a 5 x 10 array. Ideal for use with Datamouse Pro (MD7-13) canbe found on p57.Cryocanes aluminiumCryogenic vial holderThese canes hold 5 vials (290 mm long) or 6 vials (300 mm long).CryosleevesPVC sleeve to enclose CryoCane for extra security during handling andstorage. Will not become brittle while frozen.Cryocane codersUseful coloured tabs to keep track of CryoCanes. Available in white,red, green, blue and yellow. Please state colour when ordering.Cryoware labelsCloth labels for ultra low temperatures, accept ball point pen, will notshrink or peel.Cryoware marker setPack of four smudge proof extra fine point pens. Will mark cardboardand polycarbonate. Ink will not fade at ultra low temperatures. Oneeach red, green, blue and black.ORDER INFORMATIONDescription Pack CodeVial closure colour coders 100 MD7-25Cryogenic vial closure colour coders 100 MD7-26Cryogenic vial holder 1 MD7-27Cryocane for 5 vials 12 MD7-28Cryocane for 6 vials 12 MD7-29Cryosleeve 100 MD7-30Cryocane Coders 100 MD7-31(choice of white, red,green, blue & yellow)Cryoware labels 200 MD7-32Cryoware marker set 4 MD7-33114 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

BooksBOOKSA SELECTION OFCRYSTALLOGRAPHY BOOKSPractical Protein Crystallography,Duncan E. McCree, Academic Press, Second Edition, 1999.Principles of Protein X-ray Crystallography,Jan Drenth, Springer Verlag, Third Edition 2007.Macromolecular Crystallography conventionaland high-throughput methodsEdited by Mark R. Sanderson and Jane V. Skelly,Oxford University Press 2007.Protein Crystallography A Concise GuideEaton E. Lattman and Patrick J. Loll The John HopkinsUniversity Press, 2008.Protein Methods,Daniel M. Bollag, Michael D. Rozycki & Stuart J. Edelstein,Wiley-Liss, 1996.Protein Crystallization, Techniques,Strategies and Tips,Terese Bergfors, IUL, Second Edition 2008.Crystallization of Nucleic Acids and Proteins,Arnaud Ducruix & Richard Giegé, Second Edition,Oxford University Press, 1999.MD8-02MD8-04MD8-20MD8-21MD8-07MD8-08MD8-09DNA-Protein Interactions,MD8-10Andrew Travers & Malcolm Buttle, Oxford University Press, 2000.Fundamentals of Crystallography,MD8-11Giacovazzo, Monaco, Viterbo, Scordari, Gilli, Zanotti and Catti,Oxford University Press, 2000.Crystals and Life. A Personal Journey,Cele Abad-Zapatero, International University Line, 2002.Understanding Diseases byUnderstanding Proteins,Siegfried Schwarz, Karger, 2003.Introduction to Macromolecular Crystallography,Alexander McPherson, Wiley – Liss, 2008-10-09.Crystallography Made Crystal Clear,Gale Rhodes, Third Edition.MD8-12MD8-13MD8-14MD8-19Methods & Results in Crystallographyof Membrane Proteins,So Iwata, IUL, 2003.Outline of Crystallography for Biologists,David Blow, Oxford University Press, 2003.Protein Crystallization Strategies forStructural Genomics,Naomi Chayen, IUL Biotechnology Series.Biomolecular Crystallography,Bernhard Rupp, Garland Science, 2009.Textbook of Structural Biology,Liljar Anders et al, World Scientific Publishing, 2009.Membrane Protein Structure DeterminationMethods and ProtocolsLacapère, Jean-Jacques (Ed).Heterologous Expression of MembraneProteins Methods and ProtocolsMus-Veteau, Isabelle (Ed).Production of Membrane Proteins:Strategies for ExpressionRobinson, Anna Skaja (Ed).Lehringer Principles of Biochemistry (2012),Nelson, D. L. & Cox, M. M.Modelling of GPCRs,Strasser, A. & Wittmann, H-J.GPCR Signalling ComplexesDupré, D. J. Hèbert, T. E. & Jockers, R.GPCR Molecular Pharmacology andDrug TargettingGilchrist, A. (Ed).MD8-16MD8-17MD8-18MD8-22MD8-23MD8-24MD8-25MD8-26MD8-27MD8-28MD8-29MD8-30North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com115

MICROTOOLSMICROTOOLSMicrotool SetSet of eight microtools in a hardwood carrying case.Micro Scraper, Micro Knife, Micro Spade, Micro Needle, Micro Chisel,Micro Spatula, Micro Scale and Micro Prober.Can also be ordered separately.Microprobe Kit (acupuncture needles)TOOLSAN ASSORTMENT OF TOOLS FOREVERYDAY CRYSTALLOGRAPHYA useful selection of three different sizes of sterile packedacupuncture needles for crystal manipulation, tying loops, applyingsuperglue etc. Supplied in sets of thirty - ten each of three differentsizes.Measuring GridA glass measuring grid with a metallic deposited grid pattern with aline pitch of 0.1mm. Forms an ideal background on a microscopestage for measuring crystals and loops.Canned AirPressurized air to remove dust and debris from coverslips andcrystallization plates. Only available for shipping within the UK.116 Microtools117 GreasingORDER INFORMATIONDescriptionCanned AirMicroprobe Sampler KitMicro Measuring GridMicro Tool KitMicro KnifeMicro ScraperMicro NeedleMicro SpadeMicro SpatulaMicro ChiselMicro ProbeMicro ScaleMicrotool HandleFine TweezersCodeMD6-07MD9-01MD9-02MD9-03MD9-04MD9-05MD9-06MD9-07MD9-08MD9-09MD9-10MD9-11MD9-20MD9-25116 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

GREASINGGREASINGToolsGrease ApplicatorRun from your laboratory air supply, the grease applicator neatlydelivers a thread of grease as you draw it over the top of the wells ofa vapour diffusion plate.CoverSlip CombiThis single machine combines both the Grease Applicator andVacuum pen. Operated by a foot switch the machine controls bothvacuum tweezer tip for manipulating coverslips and a greaseapplicator barrel for greasing vapour diffusion plates.Finger Switch OptionThe Grease applicator, Vacuum Pick Up Pen, and CoverSlip Combimay all be ordered with a finger switch as an alternative to the footswitch.Applicator Barrel LoaderGrease ApplicatorThis useful device pressurises a tube of vacuum grease to fill thebarrels for a Grease Applicator. The Applicator Barrel Loader runsfrom your laboratory air supply and comes complete with an adaptorthat fits a grease tube into a cartridge and attaches it to the barrelfor filling. Any large pockets of air are automatically excluded fromthe grease as it is forced into the barrel.Vacuum TweezersA useful pick and place tool for manipulating coverslips.See-through pads make it easy to place coverslips accurately.Vacuum Pick Up PenAn automated version of the Tweezers, is operated by a foot switchand uses the same see-through pads as the Tweezers.CoverSlip CombiORDER INFORMATIONDescriptionPerformus VCoverslip Combi (vacuum pen and Performus V)Syringe Loader (922BL) - 2.5 OZ CartridgeFilling StationVacuum Tweezers- Hand Held Vacuum Pick Up ToolVacuum Pick Up PenGrease Applicator - Performus I F/pedalControl DispenserAir Compressor UnitTapered Tips (nozzles) - size 18 guage10ml barrels (10cc clear syringes) - White PE PistonFinger Switch- Electric10cc Air Adapter Assembly30cc Air Adapter Assembly30ml barrels (30cc clear syringes) - White PE PistonPiston U 30/55cc WH Wiper 20CodeMD9-28MD9-18MD9-19MD9-12MD9-13MD9-14MD9-26MD9-15MD9-16MD9-17MD9-21MD9-22MD9-23MD9-5112PE-BVacuum Pick Up PenNorth & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com117

GIFTWAREDECORATIVE GLASS BLOCKS OF MODEL PROTEINSAND DNA OR ORDER YOUR OWN STRUCTUREETCHED INTO GLASS FROM THE PDB CODE118 Crystal structure glass blocks118 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

GIFTWARECRYSTAL STRUCTUREGLASS BLOCKGiftwareThese decorative and impressive laser etched glass blocks make aperfect reminder of the work you have done. Give them as acongratulations gift on publication, PhD viva, leaving or retirementpresent.ORDER INFORMATIONDescriptionClassic block* 80 x 50 x 50 mmSuper size block* 100 x 70 x 65 mmGiant block* 180 x 90 x 80 mmDNA double helix 80 x 50 x 50 mmHeamoglobin 80 x 50 x 50 mmBacteriorhodopsin 80 x 50 x 50 mmImmunoglobulin G (IgG) 80 x 50 x 50 mmSuperGiant block 10 x 15 x 20 mmCube block 10 x 10 x 10 mm inc free turntableLED turntable base 110VLED turntable base 220VCodeMD14-03MD14-04MD14-05MD14-06MD14-07MD14-08MD14-09MD14-10MD14-11MD14-LED-110VMD14-LED-220V* Please email your order, complete with the PDB code forcustom blocks.All blocks include a free LED turntable base.North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com119

INCUBATORSCOMPACT, CONVENIENTBENCH TOP INCUBATORSCRYSTALLOGRAPHYINSTRUMENTATIONTemperature has always been an important factor in crystallization.Whether you're looking to simply increase the range of single pointtemperatures you routinely screen or execute ramps or cycles we havethe incubator for you. Ranging from compact space saving bench topsto large durable floor standing units, all our incubators are highlyaccurate, whilst being reliable, well made and proven to work forcrystallization. This combined with the fact they are all low or novibration (to protect the precious crystals you need to grow)makesthem the perfect choice for any lab.120 Incubators120 Bench top incubators122 Floor standing vibration freeincubators124 Liquid handling124 Optimizer126 Imaging126 Crystal X2 automated leadfinder128 X-taLight 200 automatedUV imaging systemKey Features:• Peltier-based. No compressors, no CFCs. Energy-efficient.• Temperature range from 4.0°C to 70.0°C, with an accuracy ± 0.5°C.• Choose between fully programmable or nonprogrammable controls.• 27.5, 55.0 or 100 L volumes.• Timer with alarm and Auto-Off.• Electronic calibration and powerfailure protection.• RS232 interface.• 12 month warranty.• UL, CSA and CE listed.129 X-taLight 100 UVfluorescence light source130 Dynamic light scattering130 SpectroSize 300 cuvette dls131 SpectroLight 600 sub µLscreening dls132 Pre crystallization screening133 In crystallization analysis120Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

Temperature (all units):Range 4.0° to 70.0°CReadability 0.1°CAccuracy ± 0.2°CStability ± 0.1°CSensor Type Solid State ICController Type PIDChamber Uniformity ± 0.5°CConvection Fan = YesPlate Capacity (max):SBSXRL/Linbro27.5 L model 60 2555.0 L model 140 58100.0 L model 250 112Net Weight:27.5 L unit = 16.3 kg55.0 L unit = 25.3 kg100.0 L unit = 28.0 kgElectrical:Heater/Chiller Peltier, 100 wattsPower Consumption 230 wattsElectrical 95–265 VAC input to universal power supply.12 VDC output supplied to the incubator.Power Cord Detachable, 3-wire, 2.1 meterDimensions:600/601 602 / 603 604/605Height,overall 15.5” (39.4 cm) 28.25” (71.75 cm) 28.25” (71.75 cm)Width,overall 22.25” (56.5 cm) 2.25” (56.5 cm) 22.25” (56.5 cm)Depth,overall 18.5” (47 cm) 18.5” (47 cm) 28.5” (72.4 cm)Height,chamber 12” (30.5 cm) 24” (61 cm) 24” (61 cm)Width,chamber 14” (35.6 cm) 14” (35.6 cm) 14” (35.6 cm)Depth,chamber 10” (25.4 cm) 10” (25.4 cm) 10” (25.4 cm)Stacking option:Two units can be stacked on top of one another using a spacer.Product code = MD5-606ORDER INFORMATIONDescription Volume CodeNon-Programmable 27.5 L MD5-600Fully Programmable 27.5 L MD5-601Non-Programmable 55 L MD5-602Fully Programmable 55 L MD5-603Non-Programmable 100.0 L MD5-604Fully Programmable 100.0 L MD5-605North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com121

INCUBATORSVIBRATION FREEFLOOR STANDING INCUBATORSMolecular Dimensions Cooled Crystallisation Incubatorsare an economical alternative to building temperaturecontrolled rooms, and provide much more precisecontrol of crystal growth temperature.Originally commissioned by a leading protein structure laboratoryin England, the manufacturers, have produced four superb incubatorcabinets for protein crystal growth. All have temperature programing,and can be further customised with accessories such as lighting,glazed door, extra shelves, instrument cable ports and humiditycontrol.These incubators need only a small floor space. They have a whiteinterior with easily-cleaned plastic-coated wire shelves, adjustable at35mm intervals to provide maximum flexibility of positioning.Temperature control is via an electronic controller with digital display,over/under temperature safety cut-outs and warning alarms.The doors are lockable and have a magnetic seal.The large work space door can be opened wide to provide full-widthaccess. (Glass doors are available as an option.) Particularly efficienttemperature insulation is ensured by high quality, CFC-freepolyurethane foam, enabling the pre-set temperature to bemaintained with minimum energy expenditure.WIDE VARIETY OF OPTIONAL EXTRASDescriptionMD5-04 MD5-03Part NumberMD5-02 MD5-01Additional shelf 3010 3010 3110 3210Additional shelffor lighting 1 x 20W – 3011 3111 3211Lighting 1 x 20W – 3021 3121 3221Key Features:• 100, 180, 250, and 390 litre models.• Temperature range 0°C to 50°C.• Temperature accuracy ± 0.5°C for precise crystal growth conditions.• Virtually vibration free for undisturbed experiments.• Easy-to-use control panel.• Small foot-print maximises precious floor space in the laboratory.Continuous light intensitycontrol for 1 x 20W – 3031 3131 3231Additional shelf forlighting 2 x 20W – – 3112 3212Lighting 2 x 20W – 3122 3222Continuous light intensitycontrol for 2 x 20W – – 3132 3232Glazed door – 3041 3141 3241Humidifier (evaporative) – 3061 3161 3261Work spaceelectrical outlet 3910 3910 3910 3910Cable port (20 mm) 3915 3915 3915 3915Cable port (45mm) 3916 3916 3916 3916Duration timer 3930 3930 3930 3930122Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

IncubatorsORDER INFORMATIONPrecise and constant temperatureTemperature is controlled electronically. A continuously-running fanguarantees high accuracy of temperature control throughout thecabinet interior and over time, according to DIN-58945.Heating and coolingThe continuously adjustable fan-mounted electrical resistanceheater has low thermal “drag”. It is controlled by a solid-state relay,avoiding steps and relay contact breaking. This design providesmaximal accuracy of temperature with “coasting” practicallyeliminated.The air-cooled refrigeration unit uses CFC-free refrigerant and has ahighly efficient heat exchanger. Solenoid valve-bypass operationensures smooth and continuous running, with minimal vibration.Easy-to-use, programable temperature controlA temperature program controller allowing up to 70 set points isfitted as standard. Programing is simple, with a clear digitaldisplay of set and actual temperature. Both real time and processtime controlled dual set-point temperature programs are available.Set point values or a freely programable ramp function canbe selected.Double security for sample and incubatorThe incubators cannot over-heat or over-cool. In the event of afailure, a standby controller with a separate sensor takes overtemperature control. Additionally an alarm is sounded.A mechanical safety switch with factory-adjusted releasetemperature (TWB DIN12880, class 2) protects the incubatoragainst incorrect adjustment of the temperature selector.Part NumberWork space MD5-04 MD5-03 MD5-02 MD5-01Standard minimumtemperature ±0°C ±0°C ±0°C ±0°CStandard maximumtemperature + 50°C + 50°C + 50°C + 50°CTemperature regulation ±0.5°C ±0.5°C ±0.5°C ±0.5°CVolume (Litres) 100 180 250 390Volume (cubic feet) 3.53 6.36 8.83 13.77Height (mm) 550 870 1230 1150Width (mm) 520 520 520 650Depth (mm) 440 440 440 555Exterior finish White-lacquered, zinc-plated sheet steelInterior surface Easy-to-clean white plasticNumber of shelves(standard configuration) 2 2 3 3Maximum loadper shelf (kg) 40 40 40 50CabinetHeight (mm) 1130 1440 1810 1725Width (mm) 610 610 610 760Depth (mm) 640 640 640 750Electrical power supplyoptions (V/Hz) 230/50 230/50 230/50 230/50Electrical power supplyoptions (V/Hz) 110/60 110/60 110/60 110/60Net weight (standardconfiguration) (kg) 60 60 75 90North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com123

OPTIMIZERAUTOMATED OPTIMIZATIONSCREEN MAKERLIQUID HANDLINGMolecular Dimensions presents Optimizer for eitherstraightforward construction of optimization screens orreformatting of crystallization screens.Specifically configured for handling solutions used in theoptimization of initial crystallization hits, Optimizer can rapidlyconstruct screens into either plates or deep well blocks usingsimple Excel ® files.Accurate and repeatable dispensing with low purchase andrunning costs – Optimizer can do the work for you!Key Features:• Hands-free preparation of simple Optimization Screens.• Flexible platform allowing up to 12 stock solutions or4 plates/blocks.• Rapidly re-format commercial 96 condition screens into platesor deep well blocks.• Customizable modular tube/bottle holders to suit individuallab needs.• Simple, friendly, Graphical User Interface.124Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

Optimizer can interactively display solution mixturesboth numerically and graphically for fine tuning.Working Space Dimensions:400 mm x 600 mm x 150 mm.Workspace Capacity:Adaptable with up to 4 Microplates or Deep Well blocks, 96 tubesor 48 stock solutions.Tube Holders:Available for most standard laboratory tubes, customizedformats available.Speed of motion:at least 30 cm/s.Accuracy of motion:100 µm.Pipettes:2 independent high accuracy pipettes covering volumerange of 5-200 µL and 200-1000 µL.Tips:Robotic P200 or P1000 laboratory disposable tips.Dispensing Volume:10 to 1000 µL.Dispensing Accuracy:CV from 0.2 to 2% (+/-1); max CV of 5%.Sensors:Automatic ultrasonic detection of liquid levels.ORDER INFORMATIONDescriptionOptimizerCodeMD25-01North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com125

CRYSTAL X2AUTOMATED CRYSTALLIZATIONLEAD FINDERIMAGINGThe Crystal X2 utilizes a trace label fluorescence technique toincrease both the efficiency and number of crystallization hitsidentified. Using a tiny amount of a covalent label (≤ 1%) thefluorescence can be tracked to areas in the drop where there isonly protein. Salt crystals are not “seen” and consequently thereare no false negatives as is observed when relying on intrinsictryptophan fluorescence. Plate screening is significantly speededup as intensity (generated by the harmless green LED lighting) isa faster search parameter than visible light images. Uniquely twodifferent labels can be applied to different parts of a complex toconfirm that the complex is intact in the crystal before selectingone for diffraction.Best of all, at the concentrations used, the label does notinterfere with the crystallization process or have any as yetdetectable impact on the diffraction quality. The Crystal X2 – thesimple, fast and safe protein crystallization lead finder …Key Features:• Stunning signal-to-noise ratio yields significantly highercrystallization identification hit rate.• Simple, fast and flexible labelling kit (MD1-73) 0.02% label hasno proven effect on the protein, crystallization or data quality.• Label two components with different colour dyes - uniquelycheck both parts of a complex are in the crystal.• No false positives or negatives unlike other salt/proteindifferentiation techniques.• Harmless LEDs - no risk of damage to you or the crystals.Thumbnail images display in less than 4 seconds per drop.Rapid visual identification of potential crystallization leads.126Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

Overview of Crystal X2 (MD5-700):• Provides fully automated white light and/or multiplefluorescence images.• Compatible with all SBS crystallization plates.• Operating range of 4-30 °C.• Long lifetime LEDs (50,000 hours).• Simple easy-to-use software included.ORDER INFORMATIONDescriptionCrystal X2CodeMD5-700Examples of visible (right) and fluorescentimages (left) acquired using the Crystal X2.Fluorescent labelling kits can be found on p35 of this catalogueas product codes MD1-72 and MD1-73.The presence of both parts of a complex can be confirmed using different wavelengths and dual labelling kits.North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com127

IMAGINGAUTOMATED UV IMAGING SYSTEMX-TALIGHT 200A COMBINED UV/VISIBLELIGHT BASED SINGLE PLATEIMAGING SYSTEMUtilize the intrinsic fluorescence of proteinsEvaluation is based on tryptophan fluorescence. When excitedwith light at around 300nm wavelength, it shows a strongemission at about 350nm. X-taLight allows the mixing ofUV and white light illumination for optimum image quality.• Visualize small crystals against a background of precipitate.• Minimum UV exposure time to prevent crystal damage.A fully automated imaging system, the X-talight 210 utilisesUV and white light sources to allow:• Rapid imaging and recording of crystallization experiments.• Clear differentiation of protein/other biomolecules from saltcrystals.• Visualize small crystals against a background of precipitate.• Reduce time wasted on false hits.• Fully automated CCD camera for high throughput screeningof most multi-well plates.• Minimum UV exposure time to prevent crystal damage.• 5 magnification steps: 0.63, 1.25, 2, 3.2, 6.4.• Field of view: 10.5x7.6 mm, 5.2x2.9 mm, 3.3x2.5 mm,2.0x1.5 mm and 1.0x0.75 mm.• Resolution(per pixel) – 20 µm, 13 µm, 8 µm, 5 µm and 2.5 µm.• CCD Colour Camera – 1600x1200 pixels.• Built-in temperature controller – range 4°C to 40°C (at anambient temperature of 20°C).ORDER INFORMATIONDescriptionX-taLight 200Visible Light Imaging systemX-taLight 210UV and Visible Light Imaging systemCodeMD5-200MD5-210128Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

X-TALIGHT 100OFFERS A UV FLUORESCENCESOURCE FOR YOUR EXISTINGMICROSCOPEA cost-effective, robust and easy-to-use Light Source, X-talight100 utilises integrated UV and white light sources to allow:• Clear differentiation of protein/other biomolecules fromsalt crystals.• Direct attachment to an existing laboratory microscope.• Can be upgraded to SpectroLight 600.Effectively prioritize crystallization screen hits for either follow upor diffraction. Observe directly down the microscope or captureand display images with an optional CCD camera.• Reduce time wasted on false hits.• Easily combine light sources – change the ratio of white toUV light to assess potential hits.UpgradableAs part of the modular SpectroLight system, X-taLight 100 isupgradable to a fully automated imaging system. State-of-the-art“in-drop” dynamic light scattering can also be added now or in thefuture. Be able to monitor every drop for nucleation and growth.XtaLight and SpectroLightproduced byXtal Concepts GmbHORDER INFORMATIONDescriptionX-taLight 100C Mount Microscope CameraCodeMD5-102MD5-102CCDProtein CrystalsSalt CrystalNorth & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com129

DYNAMIC LIGHTSCATTERINGGO BEYOND THE VISIBLEEnsure sample purity and qualityCheck the size distribution of your samples in the range of 1nm toseveral µm across a wide concentration range. Only small samplevolumes 8µl are needed for a complete sample characterizationwithin minutes.• Rapidly identify samples with large amounts of impurities ornonspecific aggregates (polydisperse).• Determine the dimensions and shape (when combined with SLS)of your protein as well as probing its solution state(mono/oligomer).• Examine buffer effects (e.g. salt, pH) on mode of aggregation,folding and solubility.• Analyse the effect of ligand interactions with compounds or otherbiomolecules (e.g. RNA/DNA/biopolymers/nanoparticles).SPECTROSIZE 300GIVE YOURSELF THE BESTPOSSIBLE START IN THECRYSTALLIZATION PROCESS!Multi FunctionalThe SpectroSize 300 not only allows rapid sample verification, butcan also perform more in-depth analytical tasks and long-termmeasurements including scattering angle and temperaturevariations using predefined profiles. The scattering angle can bevaried between 30 and 150° with controlled temperature variationfrom 4 to 90°C. In addition, static light scattering (SLS)measurements can be performed with ease.The SpectroSize 300 gives valuable insight into sample integrityprior to starting the crystallization process. A well defined singlespecies is widely recognised as being the key to success.SpectroSize 300 offers a rapid, simple and non-invasive solutionto understanding sample composition:• Study sample homogeneity, oligomerisation and assemblystate.• Analyse solubility, aggregation and folding effects.• Determine hydrodynamic size and mass forbiomolecules/polymers.ORDER INFORMATIONDescriptionSpectrosize 300 Cuvette Based DLSSpectroLight 600 Scanning DLSand White Light Imaging SystemSpectroLight 600 Scanning DLSand UV/White Light Imaging SystemCodeMD5-300MD5-610MD5-620130Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

SPECTROLIGHT 600SUB µL DLS FOR RAPID PRE CRYSTALLIZATIONSCREENING AND ACHIEVING DIFFICULT PROJECTSXtaLight and Spectrolightproduced byXtal Concepts GmbHThe SpectroLight range of UV/DLS instruments bringstogether the advantages of sub µl Dynamic Light Scattering andUV illumination in everyday crystallization plates. It offers rapidand powerful pre-crystallization screening of sample quality aswell as analysis of nucleation and growth throughoutcrystallization itself, and also yields informative data from everycrystallization drop, be it a clear drop or a crystal needingprotein/salt differentiation.Sub µl in plate Dynamic light scattering• Quickly screen sample stability and quality against buffers,ligands and even detergents prior to crystallization of nanolitrescale drops – saving time and sample.• Use SBS 96-well crystallization plates to analyse samplebehaviour prior to and during the crystallization experiment.• Rapidly assess homogeneity, oligomerization and assemblystate.• Track each drop's progress towards crystallization in situ andnon-invasively.• See nucleation long before crystals appear.• Rapidly record UV and white light images of experiments.• Differentiate protein/other biomolecules from salt crystals.• Visualize small crystals against a background of precipitate.How can DLS Guide Crystallization?Single narrow profile indicates lowpolydispersity – good start point!Slow nucleation – good!Nucleation too fast – shower ofmicro crystals at bestLarge scale aggregatesformed – no crystals!North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com131

DYNAMIC LIGHTSCATTERINGAPPLICATION OF SPECTROLIGHT 600IN PROTEIN CRYSTALLOGRAPHYPRE CRYSTALLIZATIONSCREENINGQUICKLY SCREEN SAMPLE STABILITYAND QUALITY AGAINST BUFFERS,LIGANDS AND EVEN DETERGENTS…HomogeneityDynamic light scattering (DLS) measurements were taken of aprotein prior to attempting crystallization. The samples wereclearly not homogeneous (Fig. 1). Attempts were made toobtain a homogeneous sample in the first case by first bufferoptimization and centrifugation (Fig.1.b).Buffer optimization: addition of 0.25 M Mg(OCO)2, incubation at4°C for 12h, clarification at 14,000 x g, adjustment of proteinconcentration to same as before addition of 0.25 M Mg(OCO)2.measurement. Typically data was recorded for one detergent at atime at a fixed time point, hence very time consuming.Using SpectroLight technology, DLS data can be rapidly obtainedfrom samples in a 96-well crystallization plate requiring only a fewmicrograms of protein per measurement and enabling severaldetergents to be screened simultaneously. In the example below, atetraspanin membrane protein was solubilised in differentdetergents at 5 x cmc (Fig. 2). Monomeric species were seen indetergents including octyl-β-D-glucopyranoside and dimers innonyl-β-Dglucopyranoside by DLS (Fig. 2.b & c), a result confirmedby SEC (Fig. 2.a).Data kindly provided byBonander, N and Oberthür, D.Figure 2: Multiple detergentsand detergent concentrationsscreened in a96-well MRC plate.Fig 2.a.Fig 2.b.Fig 2.c.Fig 1a. Heterogeneousprotein before optimization.Fig 1b. Homogeneousprotein after optimization.MonodispersityDLS of detergent-solubilised membrane proteinsin a 96-well format.The size of detergent-solubilised membrane proteins hastypically, up until now, been assessed by DLS and/or sizeexclusion chromatography (SEC) using samples of 20-100µl,which consumes tens/hundreds of micrograms of protein perStabilityIn this example (Fig. 3) the size (dynamic radius) increases over time.Hence the protein is not stable andaggregates are developing, probablyby oxidation. Often DLSmeasurements made after sizeexclusion chromatography do nottake into consideration stability overtime. With the SpectroLight 600sample stability can be monitoredeasily over hours or even days.Fig 3.132Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

IN CRYSTALLIZATIONANALYSISSTUDIES IN CONTROLLINGCRYSTAL GROWTH THROUGH DROPMONITORING AND MANAGEMENTDirecting the crystallization of ribosomeinactivating protein IIThe chosen precipitant is the right candidate for RIP IIcrystallization as indicated by the second nucleation peak.However, with a concentration of 1.8 M, the nucleation is toofast for crystallization leading to amorphous precipitation. With0.85 and 0.75 M ammonium sulphate, the nucleation peaks growmore slowly and crystals appear.At 0.75 M ammonium sulphate, the intensity of the nucleationsignal is reduced which leads to slightly bigger crystals. Hencethe optimum precipitant concentration lays somewhere between0.0 and 0.75 M.RIP II (1.2 mg/ml, approx. 60 kDa) with in situ DLS on hangingdrop crystallization experiments.RIP II under steady state conditions (A), with 1.5 M ammoniumsulphate as reservoir, mixed 1:1 with the protein drop (B), with1.7 M ammonium sulphate as reservoir (C) and with 3.6 Mammonium sulphate in the reservoir (D). Initial drop volume:2 µl.A 0.0 MB 0.75 M C 0.85 M D >1.8 MAmmonium sulphate concentration. The lower set of images are of the initial drops. Images provided courtesy of University of Hamburg.North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com133

PRODUCT INDEXDescription Page Description Page1,2,3-Heptanetriol 491,2-Propanediol 391,2-Propanediol 431,2-Propanediol 461,3-Propanediol 391,4-Butanediol 391,4-Dioxane 391,6-hexanediol 401,6-hexanediol 461.3-Propanediol 461.4-Butanediol 4610cc Air Adapter Assembly 11710 ml barrels (10cc clear syringes) - White PE Piston 1171,2,3-heptanetriol 4018 mm round no. 2 glass cover slips 6218 mm round plain coverslips No 3 glass 621-Butanol 391-Butanol 461-Propanol 3922mm round coverslips No 2 glass 6222mm round plain glass coverslips No 3 glass 6222mm square coverslips No 2 glass 622-Mercaptoethanol 502-methyl-2,4-pentanediol (MPD) 402-methyl-2,4-pentanediol (MPD) 432-Propanol 392-Propanol 4630cc Air Adapter Assembly 11730 ml barrels (30cc clear syringes) - White PE Piston 1173D Structure Screen 75 shelf rack for CXR500 Dry Shipper 11196 Square Well Storage Block Mat 7396 Well Flat Bottom Plates 72AAB buffer pH4 and pH9 45Acrylic acid/maleic acid copolymer (50:50) Na salt 38Acrylic acid/maleic acid copolymer (50:50) Na salt 48ADA 44Additional Shelf for 180L Incubator 122Additional Shelf for 250L Incubator 122Additional Shelf for 390L Incubator 122Additive Screen 36Air Compressor Unit 117Ammonium acetate 41Ammonium acetate 44Ammonium acetate 47Ammonium chloride 41Ammonium citrate 41Ammonium dihydrogen phosphate 41Ammonium formate 41Ammonium hydrogen citrate -dibasic 41Ammonium hydrogen phosphate -dibasic 41Ammonium nitrate 41Ammonium sulfate 41Ammonium sulfate 46Anzergent 3-12 49APF LB Broth Lennox 81APF LB Broth Luria 81APF LB Broth Miller 81APF Media Optimization Kit 80Assorted wicks 97Atholate 85Augmedia powder 84Bacteriorhodopsin (inc. free turntable base) 119Barium chloride dihydrate 41Bench Top Incubator 100 L Programmable 120Bench Top Incubator 100 L Non-Programmable 120Bench Top Incubator 27.5L Non-Programmable 120Bench Top Incubator 27.5L Programmable 120Bench Top Incubator 55L Non- Programmable 120Bench Top Incubator 55L Programmable 120Bench Top Incubator Stacker 121Benzamidine HCl 49Bicine 44Biomolecular Crystallography 115Bis Tris 44Bis Tris Propane 44Bovine Pituitary Extract 93BRFF-BMZERO 88BRFF-EPM2 89BRFF-HPC-1 serum free medium 89BRFF-P4-8F 90Cadmium chloride dihydrate 41Cadmium sulfate 8/3 hydrate 41Calcium acetate 41Calcium chloride dihydrate 41Calcium chloride dihydrate 46Calixar Additive Kit 23Canister for XT-20 7 XT-10 dewar 111Canned Air 116CAPS 44CAPSO 44Cesium chloride 41Cetyltrimethylammonium bromide (CTAB) 38CHAPS 49CHC buffer pH4 and pH10 45C-Hega-11 49CHES 44Chromium (III) chloride hexahydrate 41134 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

Description Page Description PageCitrate 44Classic 25 (LD Series 25L capacity) 111Classic Block (inc. free turntable base) 119Clear Sealing tape (50mm x 66M) 73Clear Strategy Screen Combination 8Clear Strategy Screen Combination pH Premixed 8Clear Strategy Screen I 8Clear Strategy Screen II 8ClearVue Sheets 73Coarse wicks 97Cobalt chloride 41Conc. HCl for titration 43Containerless Crystallization oils 51Cooled Crystallization Incubator 100 litre 122Cooled Crystallization Incubator 180 litre 122Cooled Crystallization Incubator 250 litre 122Cooled Crystallization Incubator 390 litre 122Copper(II) Chloride 41Coverslide Combi (vacuum pen and Performus V) 117Cryocane Coders 114Cryocane for 5 vials 114Cryocane for 6 vials 114CryoCap Data Matrix Reader with Vial Rack 105CryoCaps 104CryoCaps with Data Matrix 104Cryogenic vial closure colour coders 114Cryogenic vial holder 114CryoMount Elliptical LithoLoop Set 103CryoMount LithoLoop Set 103CryoMount LithoLoop Set Sampler Pack 103CryoMount Mesh LithoLoop Set 103CryoPins 18mm 104CryoProtX 43CryoProtX 109CryoProtX Mix Reagents 109Cryosleeve 114CryoTongs 24 mm 63CryoTongs for SPINE caps 112Cryotool Set 112Cryoware labels 114Cryoware Marker Set/4 Cols 114Crystal & Life Abad-Zapatero 115Crystal Quick X-ray plates 69Crystal X2 126CrystalClene Slips 18mm round 62CrystalClene Slips 18mm square 62CrystalClene Slips 22mm square 62CrystalClene Slips 22mm round 62CrystalHarp 63CrystalHarp Capillary Cutting Tool 63CrystalHarp Capillary Tweezers- rubber tipped 63CrystalHarp Starter Pack 63Crystallization of Nucleic Acids and Proteins 115Crystallography Made Crystal Clear 115Cube block 10x10x10 inc free turntable 119Custom LithoLoop CryoMount Set 103Custom screen 30CX100 Dewar lid 110CX100 Dry shipper 111CX100 Shipping Case 110CXR100 Cryogenic Shipper with Replaceable Absorbant 111CXR100 Replaceable adsorbent set 111CXR500 Dry Shipper 111CXR500 Shipping Case 111Cyclodextrin Screening Kit 96Cymal 1 49Cymal 2 49Cymal 4 49Cymal 5 49Cymal 6 49Cymal 7 49Data Logger for CX100 Dry shipper 110Data logger for CXR500 Taylor-Wharton dry shippers 111David Blow: Outline of Cryst. for Biologists 115DDM n-Dodecyl-ß-D-maltoside 49Deoxy-BigCHAP 49Detergent Screening Kit 95Deuterium Oxide 50Dewar Full Base Low Form Full Aluminum 110Dewar Full Base Low Form Full Aluminum hemispherical 110Dewar Full Base w/Metal Handle 110Dewar low form full base cylindrical 110D-galactose 46D-glucose 46Di-ethylene glycol 40Di-ethylene glycol 43Di-ethylene glycol 46Dimethyl sulfoxide 40Dimethyl sulfoxide 43DL-alanine 47DL-glutamic acid 47DL-serine 47DM (n-decyl-beta-D-maltopyranoside) 49D-mannose 47DMEM / F12 Serum-Free Medium 92DMG (n-dodecyl-NN-dimethylglycine) 50DNA Double helix (inc. free turntable base) 119DNA Protein Interactions - paperback 115North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com135

PRODUCT INDEXDescription Page Description PageDNG (decyl maltose neopentyl glycol) 49DSM (n-decyl-beta-D-thiomaltopyranoside) 49DTT 50D-xylose 47EasySeal Sheets 73EDTA 49EGTA 49Empty 96-deep well blocks 73EPPS 44ESRF Sample Changer Basket 112ESRF Sample Changer Basket tongs 112ESRF Sample Changer Canister 112ESRF Sample Changer Canister Plug 112ESRF Sample Changer COMPLETE Kit 112ESRF Sample Changer COMPLETE Kit & CryoCapsand CryoVials 112ESRF Sample Changer STARTER Kit 112ESRF Sample Changer STARTER Kit with CryoCaps & Vials 112ESRF Sample Changer Transfer Support 112ESRF Sample Changer Vial Tongs 90 deg 112Ethanol 39Ethylene glycol 40Ethylene glycol 43ExpressMAX Media Screening Kit 86F24 X-Chip Set 69F24 X-Chip Starter Kit 69F6 X-Chip Set 69F6 X-Chip Starter Kit 69Fine Tweezers 31Fine Tweezers 116Fine wicks 97Finger Switch- Electric 117Fluorescent 'Green Screen' Dye 20X stock 43FNC Coating Mix 91Foam Dewar Value Pack (1 small 1 std 1 large dewar) 110Foam Dewar Value Pack Large 110Foscarnet (phosphoformic acid) 40Foscarnet (phosphoformic acid) 50Fos-Choline 12 49Fos-Choline 9 49Freezing Media Pair 91Fundamentals of Crystallography- Giacvazzo paperback 115Gadolinium(III) chloride hexahydrate 50GeBAflex tubes 97Gelled Surface Kit 51Giant block (inc. free turntable base) 20 x 10 x 10 119Glass Capillary Tubes Borosilicate 97Glazed Panel Door for 180L Incubator 122Glazed Panel Door for 250L incubator 122Glazed Panel Door for 390L incubator 122Glucose free nutrient mix for SelenoMet media 87Glucose M9Y 84Glucose Nutrient Mix 84Glutaric acid 40Glutaric acid 50Glycerol 40Glycerol 43Glycerol ethoxylate 38Glycerol ethoxylate 48Glycine 40Glycine 44Gly-Gly-Gly 50GPCR Molecular Pharmacology and Drug Targeting 115GPCR Signalling Complexes 115Grease Applicator - Performus I F/pedal Control Dispenser 117Greiner pre-greased 24 well Combo Plate with Lid 61Greiner 24 well Combo Plate with Lid 61Hanging-drop Plate & Seal Pack 72Hanging-drop Starter Kit + JCSG+ HT-96 72Hanging-drop Starter Kit + MemGold HT-96 72Hanging-drop Starter Kit + MemStart & MemSys HT-96 72Hanging-drop Starter Kit + PACT HT-96 72Hanging-drop Starter Kit + ProPlex HT-96 72Hanging-drop Starter Kit + Structure Screen I & II HT-96 72HC35 roller base 111HC20 High Capacity Dewar 111HC34 High Capacity Liquid Nitrogen Storage Dewar 111HC35 ESRF Basket Storage System 111HC35 ESRF Canister Storage Canister 111HC35 High Capacity Liquid Nitrogen Storage Dewar 111Heamoglobin (inc. free turntable base) 119Heavy & Light Twin Pack Single Reagent 7Heavy + Light Twin pack HT-96 7Hega 10 49Hega 11 49HELIX 25HEPES 44HEPES Buffered Saline 92Heterologous Expression of Membrane ProteinsMethods and Pro 115Hexaminecobalt(III) chloride 41HTG (n-heptyl-beta-D-thioglucopyranoside) 50Humidifier - evaporative 122Hyper Broth 83I3C Phasing Kit 31IMDM Serum-Free Medium 93Imidazole 44Imidazole / DL Malic acid 45136 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

Description Page Description PageImmunoglobulin G (inc. free turntable base) 119Introduction to Macromolecular Crystallography 2nd Edition115JCSG plus 9JCSG plus Screen Single Reagent 9Jeffamine D2000 39Jeffamine ED-2001 39Jeffamine ED2003 39Jeffamine ED900 39Jeffamine M2005 39Jeffamine M2070 39Jeffamine M600 39Jeffamine SD2001 39Jeffamine T403 39KMES 44L-glutathiione 40Laminex Film Cover 73Laminex Frames 73Laminex Glass Base 100 micron 73Laminex Glass Base 200 micron 73Laminex Glass Base SBS 100 micron 73Laminex Glass Base SBS 200 micron 73Laminex Glass Cover 73Laminex Plastic Base 100 micron 73Laminex Plastic Base 200 micron 73Laminex Plastic Cover 73Laminex Starter Kit 100 micron 73Laminex Starter Kit 200 micron 73LAPAO (3-laurylamido-NN'-dimethylpropyl amino oxide) 50Large Foam Dewar 110L-arginine 40LB Booster 85LB Broth Lennox 81LB Luria Broth 81LB Miller Broth 81LD10 cryogenic dewar (20 L capacity] 111LD4 cryogenic dewar (4L capacity) 111LDAO (n-dodecyl-NN-dimethylamine-N-oxide) 49LED Turntable Light Base 110 V 119LED Turntable Light Base 220V 119L-fucose 46L-glutathione reduced 50Lighting 1 x 20 Watt for 250L incubator 122Lighting 1 x 20 Watt for 390L incubator 122Lipidic-Cubic Phase Mixing Adapter 97Lipidic-Sponge Phase Reservoir Solution 22Lipidic-Sponge Phase Screen 22Lipidic-Sponge Phase Single Reagent 22Lithium acetate 41Lithium chloride 41Lithium citrate tetrahydrate 41Lithium formate 41Lithium sulfate monohydrate 41LNG (lauryl maltose neopentyl glycol) 49Low Level Alarm for HC34 dewar 111Low Liquid Alarm for HC35 VHC35 XT20 111Low Profile Laminex Frames 73L-proline 40LS Series 35L Liq N2 storage system for racks(750 samples) 111Lysine HCl 40Lysine HCl 47Macromolecular Crystallography- Conventional and HTmethods 115MacroSol 14MacroSol Screen Single Reagent 14Magnesium acetate 41Magnesium chloride hexahydrate 41Magnesium chloride hexahydrate 46Magnesium formate dihydrate 41Magnesium nitrate hexahydrate 41Magnesium sulphate 41Magnetic 45 degree bent angle Cryo Wand for SPINE caps 112Magnetic Cryo Wand for SPINE caps with Suregrip handle 112Magnetic CryoVial 104Magnetic CryoVial with CryoCaps 104Magnetic CryoVial with CryoCaps with data matrix 104Maltose 43Manganese(II) chloride tetrahydrate 50Media Optimization Kit 81Medium wicks 97MemAdvantage 24Membrane Protein Combination 17Membrane Protein Structure DeterminationMethods and Protocol 115MemGold 19MemGold Combo Value Pack 19MemGold Single Reagent 19MemGold2 19MemGold2 Single Reagent 19MemMagic Bicelle Screen Kit 23MemPlus 18MemPlus Screen Single Reagent 18MemStart + MemSys 17MemStart 17MemStart Single Reagent 17MemSys 18MemSys Single Reagent 18MERPOL HCS 49North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com137

PRODUCT INDEXDescription Page Description PageMES 44MES/Bis Tris 44Methanol 39Methionine solution 250x 87Methods & Results in Cryst. of Membrane Proteins,So Iwata 115MIB pH4 and pH10 45Micro Chisel 116Micro Knife 116Micro Measuring Grid 116Micro Needle 116Micro Probe 116Micro Scale 116Micro Scraper 116Micro Spade 116Micro Spatula 116Micro Tool Kit 116Microprobe Sampler Kit 31Microprobe Sampler Kit 116MicroSeed Beads 31Microspatula 31Microtool Handle 116MIDAS 15MIDAS OptiMax Kit 15MIDAS Single Reagent 15Mineral Oil 43Mini Screen 7Mini Screen Single Reagent 7MMT pH4 and pH9 45Modelling of GPCRs - 2013 115Monoolein 22MOPS 44Morpheus 13Morpheus - Buffer System 1 45Morpheus - Buffer System 2 45Morpheus - Buffer System 3 45Morpheus Additive OptiMax Kit 13Morpheus- Alcohols Mix 45Morpheus- Amino Acids Mix 45Morpheus- Buffer 1 Mix 45Morpheus- Buffer 2 Mix 45Morpheus- Buffer 3 Mix 45Morpheus- Carboxylic Acids Mix 45Morpheus- Divalents Mix 45Morpheus- EDO_P8K Mix 45Morpheus- Ethylene Glycols Mix 45Morpheus- GOL_P4K Mix 45Morpheus- Halogens Mix 45Morpheus- Monosaccharides Mix 45Morpheus- MPD_P1KP3350 Mix 45Morpheus- NPS Mix 45Morpheus- P550MME_P20K Mix 45Morpheus Screen Single Reagent 13Mounted Elliptical LithoLoops 102Mounted LithoLoop Sampler Pack (NEW) 102Mounted LithoLoop Sampler Pack-CUSTOM 102Mounted LithoLoops 102Mounted Mesh LithoLoops 102MRC Crystallisation Plate Polystyrene 68MRC Crystallisation Plate UV 68MRC MAXI Optimization Plate (Polystyrene) 68MRC MAXI Optimization Plate (UV) 68MRC Under Oil Crystallization Plate - UVP 69MultiXtal 7Na/K phosphate 44N-acetyl-D-glucosamine 46Naomi's Nucleants 31NDSB-201 43NG (n-nonyl-beta-D-glycopyranoside) 49Nickel chloride 41Nickel II sulfate hexahydrate 41NM (n-nonyl-beta-D-maltopyranoside) 49Notched metal tubes 104Nozzle allignment tool 105NR-LBD + NR-LBD Extension Screen Single Reagent 16NR-LBD Extension Screen 16NR-LBD Extension Screen Single Reagent 16NR-LBD Screen 16NR-LBD Screen Single Reagent 16Nuclear Receptor Combination 16Octaethylene glycol monododecyl ether (C12E8) 50OG (n-octyl-β-D-glucoside) 49OM-fluorinated (octyl maltoside flourinated) 49ONG (octyl glucose neopentyl glycol) 49OptiClear Seals for MRC Crystallization Plates DLS 73Optimizer 125Osmium(III) chloride hydrate 50PACT premier 9PACT premier Screen Single Reagent 9Paraffin oil 51Parrafin oil 43PCTP pH4 and pH9.5 45PEG 1000 37PEG 10000 37PEG 12000 37PEG 1500 37PEG 200 37PEG 2000 37138 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

Description Page Description PagePEG 2000 MME 37PEG 20000 37PEG 300 37PEG 3000 37PEG 3350 37PEG 350 MME 37PEG 400 37PEG 4000 37PEG 500 MME 37PEG 5000 MME 37PEG 600 37PEG 6000 37PEG 750 MME 37PEG 8000 37Penta erythritol propoxylate (5/4 PO/OH) 48Pentaerythritol ethoxylate (15/4 EO/OH) 38Pentaerythritol ethoxylate (15/4 EO/OH) 48Pentaerythritol ethoxylate (3/4 EO/OH) 48Pentaerythritol ethoxylate (3/4 EO OH) 38Pentaerythritol propoxylate (17/8 PO/OH) 38Pentaerythritol propoxylate (17/8 PO/OH) 48Pentaerythritol propoxylate (5/4 PO/OH) 38Penta-ethylene glycol 40Penta-ethylene glycol 46Performus V 117PERK - Protein Expression Rescue Kit 85PET: Cell Dissociation Formula 90PGA Screen 22PGA screen Single Reagent 22PGA-HM 38Phosphate/citrate buffer 44PIPES 44Piston U 30/55cc WH Wiper 20 117Poly Carb Box for CXR500 Dry Shipper 111Polyacrylate 2 sodium salt 48Polyacrylate 2100 38Polyacrylate 5 sodium salt 48Polyacrylate 5100 38Polyacrylic acid 5100 38Polyethyleneimine 38Polyethyleneimine 48Poly-glutamic acid (PGA-LM) 38Polypropylene glycol 400 38Polypropylene glycol 400 48Polyvinyl alcohol type II 38Polyvinyl alcohol type II 48Polyvinylpyrrolidone (PVP) 38Polyvinylpyrrolidone (PVP) 40Potassium acetate 41Potassium bromide 41Potassium chloride 42Potassium citrate 44Potassium di-hydrogen phosphate (monobasic) 42Potassium fluoride 42Potassium fluoride 50Potassium formate 42Potassium hydrogen phosphate (dibasic) 42Potassium iodide 42Potassium nitrate 42Potassium phosphate 44Potassium silicate 42Potassium silicate 50Potassium sodium tartrate 42Potassium thiocyanate 42Power Broth 83Power Combo Value Pack (Morpheus & MIDAS) 13Power Prime Broth 83Practical Protein Crystallography 115Prime-olate media optimization kit 81Principles of Biochemsitry -2012 115Principles of Protein X-Ray Crystallography 3rd Ed 115Production of Membrane Proteins: Strategiesfor Expression 115ProPlex 15ProPlex Single Reagent 15Protein Crystallisation Techniques Strategies & Tips 115Protein Crystallization Strategies for Structural Genomics 115Protein Crystallography: A Concise Guide 115Protein Methods 2nd Ed 1996 115QuickFold Protein Refolding Kit 95Really Useful Buffer Kit 45Really Useful Buffer Kit 51Roller 72Roller Base for Classic 25 (LD Series 25L capacity) 111Roller Base for XT-20 Liquid Nitrogen Storage Dewar 111Roller for LS Series 35L Liq N2 Storage system for racks 111Rotating Light Base (LUMINORUM) 119Rubidium chloride 42Rubidium chloride 50Samarium(III) chloride 50Sealing grease 61Sealing Sheet Applicator 73SelenoMet base media and nutrients 87SelenoMet base medium and Nutients Glucose Free 87SelenoMet Medium Base 87SelenoMet Medium Complete 87SelenoMet Medium Complete Glucose-Free 87SelenoMethionine Solution 87North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com139

PRODUCT INDEXDescription Page Description PageSeMet Nutrient Mix 87Serum-Free Medium Screening Kit 94Si coverslips 18mm round 62Si coverslips 22mm round 62Si coverslips 22mm square 62SILAC- L-Arg 87SILAC- L-Leu 87SILAC- DMEM high glucose media 87SILAC- DMEM low glucose media 87SILAC- DMEM/F12 87SILAC- Harm's F12 87SILAC- L-Lys 87SILAC- L-Met 87SILAC- McCoy's 5A 87SILAC- MEM 87SILAC- RPMI 1640 87Silicone oil 51Single reagents 36Sitting drop bridges 61Small Foam Dewar 110Sodium acetate 42Sodium acetate 44Sodium azide 42Sodium azide 50Sodium borate 44Sodium bromide 42Sodium bromide 46Sodium cacodylate 44Sodium chloride 42Sodium citrate 42Sodium citrate 47Sodium citrate 44Sodium fluoride 42Sodium fluoride 46Sodium formate 42Sodium formate 47Sodium Hepes 44Sodium hydrogen phosphate dibasic 42Sodium hydrogen phosphate dibasic 46Sodium hydroxide 43Sodium iodide 42Sodium iodide 46Sodium malonate 42Sodium metaborate 42Sodium nitrate 42Sodium nitrate 46Sodium oxamate 42Sodium oxamate 47Sodium phosphate 44Sodium phosphate dibasic 42Sodium sulfate 42Sodium tartrate 42Sodium thiocyanate 42Sodium/Potassium phosphate (PACT) 42Sokalan CP 42 38Sokalan CP 5 38Sokalan CP 5 48Sokalan CP 7 38Sokalan CP 7 48Sokalan HP 56 38Sokalan HP 56 48Solubility Tool Kit - Precipitant Screen Box 2 8Solubility Tool Kit - Solubility Screen Box 1 8Solubility Tool Kit (Box 1 + 2) 8Solubility Tool Kit Single Reagent 8SpectroLight 600 scanning DLS and UV/White lightimaging system 130SpectroLight 600 scanning DLS and white lightimaging system 130SpectroSize 300 Cuvette based DLS instrument 130Spermidine 40Spermidine 50Spermine HCl 40SPG pH4 and pH10 45Standard Foam Dewar 110Strontium chloride 42Strontium chloride 50Structure Screen 1 5Structure Screen 1 + 2 Single Reagent 5Structure Screen 1 Single Reagent 5Structure Screen 2 6Structure Screen 2 Single Reagent 6Structure Screen Combination 5Structure Screen 1 + 2 5Stura Footprint Combination 6Stura Footprint Screen Single Reagent 6Stura Footprint screens 6Stura MacroSol Single Reagent 14Succinic acid 40Sucrose monodecanoate (sucrose monocaprate) 49Sucrosse monododecanoate 50Super size block (inc. free turntable) 119Super2 Combo Value Pack (JCSG+ & PACT) 9Superglue for CryoPins 104140 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

Description Page Description PageSuperior Broth 82Superior Prime Broth 82SureSeal DWB 73Swissci Hanging Drop Plate -UVP 72Swissci LCP Kit 73Swissci Triple Drop Plate Polystyrene 68Swissci Triple Drop Plate UV Polymer 68Syringe Loader (922BL)- 2.5 OZ Cartridge Filling Station 117Tall Foam Dewar 110Tapered Tips (nozzles)- size 18 guage 117Taurine 40Taurine 50TBG buffer pH4 and pH9 45TCEP 50Terasaki Microbatch Plate 72-well 69Tertiary-butanol 39Tetra-ethylene glycol 40Tetra-ethylene glycol 46Tetraethylene glycol monooctyl ether (C8E4) 50Tetrahydrofuran 39Textbook of Structural Biology 115Trace Fluorescence Dual-Labelling Kit 35Trace Fluorescence Labelling Kit 35Trehalose 40Trehalose 43Tri DM (n-tridecyl-beta-D-maltoside) 49Trial Pack 10 MRC Crystallization Plates 10 ClearVue Sheets 68Tricine 44Triethylammonium phosphate 50Triethylene glycol (TEG) 40Triethylene glycol (TEG) 46Trimethylamine N-oxide (TMAO) 38Tripao 49Tris 44Tris hydrochloride 44Tungsten Carbide Scriber 73Turbo Broth 82Turbo Prime Broth 82UDM (n-undecyl-beta-D-maltoside) 50UDTM (n-undecyl-B-D-thiomaltopyranoside) 50Understanding Diseases by Understanding Proteins 115UV Hanging Drop Sheets 72Vacuum Pen- Air Powered F/Pedal Control Vacuum Pen 117Vacuum Tweezers- Hand Held Vacuum Pick Up Tool 117VHC35 High capacity Liquid Nitrogen Storage Dewar 111VHC35 roller base 111Vial Tongs 45 Degrees 112Volatile Oil for MRC Under Oil Plate 51Water 18 MegaOhm purified/filtered 1 Liter 43XRL Plate 61XRL Plates incl. CrystalClene Slips 61XT-10 Extended Time Cryogenic dewar (10 L) 111XT-20 Extended Time Cryogenic dewar (20 L) 111X-taLight 100 129X-taLight 200 White light imaging system 128X-taLight 210 UV and white light imaging system 128XTL3 Extended Time Cryogenic dewar (3 L) 111Xylitol 40Xylitol 43Zinc acetate 42Zinc chloride 42Zinc nitrate hexahydrate 42Zinc nitrate hexahydrate 50Zinc sulfate heptahydrate 42We kindly acknowledge the following people for their crystalimages used throughout this publication: Alexey Rak, Karolien VanBelle, Mareike Kurz, Isabell Wingartz, Simon Newstead, MarekBrzozowsky, Gary Parkinson, Stephen Jameison, Annabelle Varrot,Natacha Perebaskine, Anwar Ullah, Karolina Mierzejewska, AlbertGuskov, Daljit Sangar and Jane Potter.North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com141

Notes142 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

North & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.com visit: www.moleculardimensions.com143

Notes144 Europe and rest of world orders: +44 (0)1638 561 051 email: orders@moleculardimensions.com visit: www.moleculardimensions.com

How to orderYou can place your order online, by fax, telephone or via e-mail. Our site includes a full e-commercefacility for selecting and ordering your product. Credit card orders can be placed securely on-line.ReturnsReturns will not be accepted without prior authorization. Damaged items must be notified within 7 days.PricesPrices are subject to change without notice. There is no minimum order charge.To place an order in Europe or the rest of the world, excluding theAmericas please place your order with our UK office.Molecular Dimensions Ltd.Unit 6, Goodwin Business Park,Willie Snaith Road, Newmarket, Suffolk, CB8 7SQ,UKTelephone: +44 (0)1638561051Fax: +44 (0)1638660674To order by e-mail useorders@moleculardimensions.comTo order on account please contact us to set up an account.ShippingProducts are shipped by DHL.To place an order in the USA, Canada, Mexico or South America pleaseplace your order with our USA office.Molecular Dimensions Inc.849 Sunshine Lane, Altamonte Springs, Florida, 32714USATelephone: +1 877 479 4339or +1 407 886 6901Fax: +1 321 972-8896To order by e-mail useusorders@moleculardimensions.comTo order on account please contact us to set up an account.ShippingProducts are shipped by Fedex.DistributorsCHINABio-Direct Co. Ltd.Room 301, Building No. 2.88 Darwin Road,Pudong, 201203Shanghai, P.R. ChinaTel: +86 21 5197 3610Beijing Seaskybio Technology Co.LtdRoom 904A Building,Maples International Centre,No. 32 Xizhimen North Street,Haidian District, Beijing, China, 100082Tel: +8610-62220605Fax: +8610-62168941INDIARFCL Limited, a part ofAvantor Performance Materials12th Floor, Pinnacle Claridges Business Tower,Shooting Range Road, Surajkund,Faridabad-121009, IndiaTel: +91-129-4267059Fax : +91-129-4267199JAPANWaken B Tech, Co., Ltd.Sales Promotion Dept.1-12-8 Senbahigashi, Minoh-city,Osaka 562-0035, JapanPhone: +81-72-749-5300Fax: +81-72-749-5600http://www.wakenbtech.co.jpKOREAMDxK Molecular Diagnostics Korea Inc.T: +82 2 578 8848F: +82 2 578 88384F Song B/D, 333-16,Yungjae-Dong, Seocho-Gu,Seoul 137-897, Korea (Republic of)MALAYSIAEko-Teknik15-01Wisma ZelanNo.1 Jalan Tasik Permaisuri 2Bandar Tun Razak56000 Kuala LumpurMALAYSIATel: +603 91736440Fax: +603 91735451RUSSIAN FEDERATIONVladimir ChernyavskyNagatinskaya Naberezhnaya 54, Suite 577Moscow, Russia 115407Tel. +79167575787Tel. +79857773773SINGAPOREPractical Mediscience Pte Ltd+65 6254 2633551-C Baletier Road, Singapore 329868,SingaporeBio-Etc Pte Ltd (Email: ngch@bio-etc.com)41 Science Park Road#04-06A The GeminiSingapore Science Park IISingapore 117610Mobile: +65 8617 7360Office: +65 6779 4149Fax: +65 6779 4095TAIWANYeong Jyi Chemical+886 (2) 2982 8796Apparatus Co. LimitedPO Box 117, Sanchung, Taipei, TaiwanNorth & South America orders: +1 877 479 4339 email: usorders@moleculardimensions.comvisit: www.moleculardimensions.com145

Molecular Dimensions Ltd.Unit 6, Goodwin Business Park,Willie Snaith Road,Newmarket, Suffolk, CB8 7SQ,UKTelephone: +44 (0)1638561051Fax: +44 (0)1638660674Molecular Dimensions Inc.849 Sunshine Lane, Altamonte Springs,Florida, 32714USATelephone: +1 877 479 4339or +1 407 886 6901Fax: +1 321 972-8896