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J Virol Methods 2006 Jul;135(1):32-42

Real-time, quantitative PCR assays for the detection of virus-specific DNA in samples with mixed populations of polyomaviruses.

Pal A, Sirota L, Maudru T, Peden K, Lewis AM Jr

Lewis AM, US FDA, CBER, Div Viral Prod, Off Vaccines Res & Review, Bldg 29A,Room 1BO13,29 Lincoln Dr, Bethesda, MD 20892 USA US FDA, CBER, Div Viral Prod, Off Vaccines Res & Review, Bethesda, MD 20892 USA US FDA, CBER, Div Biostat, Off Biostat & Epidemiol, Bethesda, MD 20892 USA


Mixtures of polyomaviruses can be present in the central nervous system, the gastrointestinal tract, the genitourinary tract, blood, and urban sewage. We have developed 12 primer/probe sets (four per virus) for real-time, quantitative PCR assays (TaqMan) that can specifically detect BKV, JCV, and SV40 genomes present in mixtures of these viruses. The specificities of these primer/probe sets were determined by evaluating their level of interaction with the DNA from other polyomaviruses and their ability to estimate the number of copies of homologous viral DNA in blinded samples of defined mixtures of three polyomaviral DNAs. Three early region and three late region primer/probe sets determined, within a two-fold range, the number of copies of their respective DNAs. Four sets of SV40 primer/probes also detected 1.1-2.4 copies of SV40 DNA per COS-1 cell, cells estimated to contain a single copy of SV40 DNA. Three JCV primer/probe sets detected 3.7-4.2 copies per cell of JCV DNA in the JCV-transformed cell line M1-HR, cells estimated to contain between 0.5 and 1 copy of the JCV genome. We suggest that the virus-specific primer/probe sets in this study be considered sufficiently characterized to initiate the quantification of polyomavirus DNA in biological samples.

Category: Journal Article
PubMed ID: #16527364
Includes FDA Authors from Scientific Area(s): Biologics
Entry Created: 2011-10-04 Entry Last Modified: 2012-08-29