Detection of HIV-1 DNA by PCR: evaluation of primer pair concordance and sensitivity of a single primer pair.
Concordance between two primer pairs and the clinical sensitivity of a single primer pair-probe system were evaluated for a human immunodeficiency virus type 1 (HIV-1) polymerase chain reaction (PCR) assay. Six-hundred sixty-three clinically defined HIV-1 specimens were analyzed in a blind fashion for HIV-1 DNA using an optimized, well-characterized PCR assay. All samples were amplified in duplicate with each of two primer pairs targeting distinct, highly conserved regions within the HIV-1 gag genome. Amplified product was detected by oligomer hybridization using 32P-labeled probes. Correlation between PCR, culture, and serology data was achieved in 661 of 663 (99.7%) specimens. After initial analysis, discordant PCR results due to discrepancies between primer pairs were observed in 9 of 663 (1.4%) specimens. All nine specimens were resolved as negative on reevaluation. After retesting of discordant PCR results, concordance between the two primer pairs was achieved in 100% of samples. These data indicate that the two primer pair systems performed comparably and that good clinical sensitivity (100%) would be achieved using a single primer-probe system. Factors and procedures that influence the reproducibility and accuracy of this HIV-1 PCR assay are discussed.
Lynch
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Roche Molecular Systems, Inc., Alameda, CA 94501.
Madej
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R
Louie
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PH
Rodgers
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D015492:AIDS Serodiagnosis / Q000379:methods; D001483:Base Sequence; D002478:Cells, Cultured; D015342:DNA Probes; D015343:DNA Probes, HLA; D004279:DNA, Viral / Q000032:analysis* / Q000737:chemistry; D005260:Female; D015497:HIV-1 / Q000235:genetics* / Q000254:growth & development; D006801:Humans; D008297:Male; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods / Q000592:standards*; D012189:Retrospective Studies
D015342:DNA Probes; D015343:DNA Probes, HLA; D004279:DNA, Viral