The reverse transcriptase assay: problems and practical solutions.
The detection of retroviruses has become critical for addressing the safety concerns associated with biologically derived products, including those derived from blood and cell line substrates, and gene therapy based systems. Most, if not all, retroviruses encode a unique enzyme called reverse transcriptase whose RNA-dependent DNA polymerase activity can be used as a marker for detecting retroviral contamination in test material. In this presentation we document some practical concerns when using the reverse transcriptase assay for detection of retroviruses. We also illustrate important aspects in the assay design by presentation of results that needed supplemental testing to enable accurate assessment of retroviral contamination.
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Quality Biotech Inc., Camden, NJ 08104, USA.
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D000818:Animals; D002460:Cell Line / Q000821:virology; D002852:Chromatography, Ion Exchange; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism; D004340:Drug Contamination; D005189:False Positive Reactions; D006801:Humans; D007700:Kinetics; D008274:Magnesium; D008345:Manganese; D012194:RNA-Directed DNA Polymerase / Q000032:analysis*; D012190:Retroviridae / Q000201:enzymology*; D013698:Templates, Genetic; D013942:Thymine Nucleotides / Q000378:metabolism; D014773:Virology / Q000379:methods*
D013942:Thymine Nucleotides; D008345:Manganese; D012194:RNA-Directed DNA Polymerase; D004259:DNA-Directed DNA Polymerase; D008274:Magnesium; C024157:thymidine 5'-triphosphate