کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
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3408465 | 1223665 | 2006 | 9 صفحه PDF | دانلود رایگان |
A sensitive and reproducible method to determine the in vitro infectious potency of a pentavalent reassortant rotavirus vaccine (RotaTeq®) has been developed as an alternative to classical potency assays. Potency was determined based on cell-based viral replication followed by quantitative reverse-transcription polymerase chain reaction (RT-QPCR) analysis. In the assay, confluent Vero cell monolayers in 96-well plates were inoculated with serial dilutions of test samples, a pentavalent reassortant rotavirus reference standard and assay controls, followed by incubation for 24 h. The cells were lysed with a Triton X-100 solution and the lysates assayed by RT-QPCR to quantitate viral nucleic acid produced during replication. The RT-QPCR utilizes primer/probe sets specific to each virus reassortant and the potencies of each sample were determined relative to the reference standard. This assay, hereafter referred to as the Multivalent QPCR-Based Potency Assay (M-QPA), permits the specific quantitation of each individual reassortant virus in the presence of the other four reassortant viruses. In addition, the assay was demonstrated to be concordant with a traditional method (plaque assay) for the quantitation of infectious virus particles. It is anticipated that assays of this type will become a valuable tool in the assignment of potency values and in the monitoring of stability of live virus vaccines.
Journal: Journal of Virological Methods - Volume 131, Issue 2, February 2006, Pages 193–201