2011-2012 Catalogue - PeproTech, Inc.

PEPROTECH ®www.peprotech.comELISA FAQ1. How many plates can I run with one ofPeproTech’s ELISA Development Kits (EDKs)?One EDK contains enough material to run 10standard plates at 100 wells each. Thiscorresponds to 1,000 ELISA plate wells.2. What is the stability of the avidin-HRPincluded in PeproTech’s ELISA Development Kit?The avidin-HRP included in the kit is stable forup to 1 month at 2-8˚C, and for up to 2 years at-20˚C.3. How can I find cross reactivity information for akit?PeproTech performs in-house cross reactivitytesting on its ELISA Development Kits. Theresults that have been collected from this testingare located on the kit’s data sheet.4. Is there any step of the ELISA protocol that canbe left over the weekend?The plate may be coated with the captureantibody on Friday, left at 4˚C over the weekend,and resumed on Monday. Please note: changingincubation times may cause results to varybetween plates.5. Are PeproTech’s EDKs suitable to use with allsample types?Although PeproTech has not tested all of its kitsin every matrix available, they should be suitablefor use in, but not limited to: serum, plasma, cellculture supernatant, urine and saliva.6. Is a stop solution necessary to stop thereaction?A stop solution is not needed when usingavidin-HRP + ABTS. In general, reliable standardcurves are obtained when either O.D. readings donot exceed 0.2 units for the zero standard concentrations,or 1.2 units for the highest standardconcentration.If a stop solution is desired, 1% sodium dodecylsulfate (SDS) may be used to end the reaction.Stop solutions are not used in PeproTech’slaboratory, though literature has shown that SDS isthe optimal stop solution for ABTS.7. Can I use TMB with PeproTech’s EDKs?PeproTech’s EDKs are optimized using ABTSand are therefore best used in conjunction withthis substrate. The kit can still be used in combinationwith TMB, but only after some adjustmentshave been made:• The avidin-HRP provided in the kit cannot beused with TMB; streptavidin must be purchasedseparately.• Dilutions of streptavidin will need to beoptimized.• A stop solution is generally needed when usingstreptavidin + TMB. Refer to manufacturer’s datasheet.• The TMB reaction time, prior to the addition ofstop solution, will need to be optimized.• The plate is to be read at 450nm with a correctionat 650nm when using recommended plates.8. Why is D-mannitol added to the EDKcomponents?D-mannitol is added to the EDK components inorder to aid in protein/antibody visualization. Itdoes not alter ELISA results.9. Can I use the curve on the EDK data sheet as mystandard curve?A separate standard curve must be run on eachELISA plate. In other words, the curve from oneplate cannot be used for a different plate. Thecurve that PeproTech provides on the EDK datasheet is for demonstration purposes only, asachieved in PeproTech’s laboratory.10. How does PeproTech generate their standardcurve?When an ELISA is run in PeproTech’s lab, aMolecular Devices ® plate reader and theSOFTmax ® PRO software are used. This programuses the values that are received and generates a4-parameter curve. Here is the equation that theprogram uses:4-P Fit:x = concentration (pg/ml)y = O.D. (405nm – 650nm)A, B, C, & D correspond to the 4 parameters.**For more detailed information regarding the parameters,please contact the Technical Support Department.INFORMATION & TECHNICAL SUPPORTBestellungen: Deutschland: Tel: 0800 436 99 10 • email: info@peprotech.dePour commander: France: Tel: +33 (0)1 46 24 58 20 • email: info@peprotechfr.comTo Order: NORDIC: Tel: +46 (0)8 640 41 07 • email: info@peprotech.se161