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genomics For more information on these products contact your local VWR sales office, send an e-mail to vwrbiomarke@eu.vwr.com or visit our website www.vwr.com Figure 2: Pilot demonstration equipment showing foam fractionation. A culture is grown in the vessel (A) and produces surfactin. Air from the sparger (B) blows foam into the J-tube (C). As the foam travels up the column, medium drains downwards by gravity, so that the ‘foamate’ collected in the beaker (D) contains 40 – 50x more surfactant than the culture broth. The ‘foamate’ also contains a small amount of biomass. Measuring mRNA levels in (A) provides information on cellular biosynthetic activity, while measuring 16S rDNA in (D) gives an accurate bacterial count of the degree of carryover. added, particularly useful given the numbers of different primers used in a single plate. Most of our work has been done using SYBR ® Green I, but we have obtained comparable results using corresponding master mixes for probe detection too. A further improvement in ease of use was the recent introduction of the white ABgene SuperPlate . We tried them to see by how much the white colour would improve assay sensitivity. We found that assay sensitivity improved even at very low template concentrations. Pipetting clear reagents into white plates is difficult, which increases the likelihood of introducing errors. This may cause more handling problems than can be justified by the increase in sensitivity. However, the blue coloured Thermo Scientific Verso QRT-PCR reagents enabled us to spot our pipetting errors before they impacted our results. Consequently, we were able to develop an extremely reliable assay. References 1. Bonmartin, JM; Laprevote, O; Peypoux, F. (2003). Diversity among cyclic lipopeptides: iturins and surfactins. Activitystructure relationships to design new bioactive agents. 2. Combinatorial Chemistry and High Throughput Screening 6 (6) 541-546. Chen, CY; Baker, SC; Darton, RC (2006). Continuous production of biosurfactant with foam fractionation. Journal of Chemical Technology & Biotechnology 81(12) 1915-1922. Description Pack Cat. No. Verso 1-Step QRT-PCR Kit plus ROX vial 400 x 25 µl 733-1114 Verso SYBR Green 1-Step QRT-PCR kit plus ROX vial 400 x 25 µl 733-1129 ABgene Superplate PCR detection plate 25 plates 732-1056 ABgene Superplate PCR detection plate, white 25 plates 731-0305 ABgene Superplate semi-skirted PCR 25 plates 732-1060 ABgene Superplate semi-skirted PCR, white 25 plates 732-1063 ABgene Superplate skirted PCR plate 25 plates 732-1049 ABgene Superplate skirted PCR plate, white 25 plates 732-1052 ABgene Superplate semi-skirted raised deck, barcoded 25 plates 732-1492 ABgene Superplate semi-skirted raised deck, barcoded, white 25 plates 732-1493 ABgene Superplate semi-skirted flat deck, barcoded 25 plates 732-1494 ABgene Superplate semi-skirted flat deck, barcoded, white 25 plates 732-1495 ABgene Superplate skirted low profile, barcoded 25 plates 732-1496 ABgene Superplate skirted low profile, barcoded, white 25 plates 732-1497 VWR International I VWRbioMarke Issue 27 I September 2011 I 11
the market source for life science DNA/RNA extraction from formalin fixed, paraffin embedded tissue Formalin Fixed, Paraffin Embedded (FFPE) tissue is one of the most widely practiced methods for clinical sample preservation and archiving. It is estimated that, worldwide, over a billion tissue samples, most of them FFPE, are being stored in numerous hospitals, tissue banks, and research laboratories. These archived samples could potentially provide a wealth of information in retrospective molecular studies of diseased tissues. While standard for histopathology and microscopic investigation (e.g., Hematoxylin and Eosin [H&E] staining, ImmunoHistoChemistry [IHC], andTissue MicroArray [TMA]), FFPE samples pose a major challenge for molecular pathologists, because nucleic acids are heavily modified and trapped by extensive proteinnucleic acid and protein-protein cross-linking. Historically, FFPE samples were not considered to be a viable source for molecular analyses. Recently, however, it has been discovered that with appropriate protease digestion, it is possible to release microgram amounts of DNA and RNA from FFPE samples. The purified nucleic acids, although highly fragmented, are suitable for a variety of downstream genomic and gene expression analyses, such as polymerase chain reaction (PCR), quantitative reverse transcription PCR (qRT-PCR), microarray, array comparative genomic hybridisation (CGH), microRNA, and methylation profiling. Omega Bio-tek offers a complete line of products for your FFPE extraction needs. The E.Z.N.A ® spin column-based kits are ideal for low throughput applications, whereas our Mag-Bind ® family of magnetic beadsbased kits are designed specifically for high throughput users with automation capability. Together, these kits provide clinical researchers a comprehensive and robust sample preparation solution for biomarker discovery, validation and application. Figure 1. Representative gel analysis of DNA extracted from FFPE liver tissue using the Mag- Bind ® FFPE DNA 96 kit (left) and E.Z.N.A. ® FFPE DNA Isolation kit (right). Figure 2. Representative Agilent Bioanalyzer trace of RNA extracted from FFPE liver tissue using the E.Z.N.A. ® FFPE RNA Isolation kit. Table 1. Yield and quality of DNA extracted from FFPE liver tissue using the E.Z.N.A. ® FFPE DNA/RNA Isolation kit (column) and Mag-Bind ® FFPE DNA/RNA 96 kit (magnetic beads) DNA RNA Method Column Beads Conc. (μg/μL) Yield (μg) 260/280 260/230 Conc. (μg/μL) Yield (µg) 260/280 260/230 0,51 5,1 1,85 1,6 0,32 44,5 2,13 2,16 0,49 4,9 1,84 1,6 0,32 44,6 2,12 2,16 0,51 5,1 1,83 1,5 0,25 34,9 2,12 2,17 0,67 6,7 1,85 1,7 0,35 34,5 2,11 1,82 0,72 7,2 1,82 1,9 0,34 34,3 2,11 1,81 0,79 7,9 1,83 1,7 0,32 32,4 2,10 1.79 12 I VWR International I VWRbioMarke Issue 27 I September 2011
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