A simple competitive RT-PCR assay for quantitation of HIV-1 subtype B and non-B RNA in plasma

Makiko Hamatake, Masako Nishizawa, Naoki Yamamoto, Shingo Kato, Wataru Sugiura

Research output: Contribution to journalArticle

5 Citations (Scopus)

Abstract

An easy, inexpensive competitive RT-PCR assay for HIV-1 RNA quantitation was constructed. A 138-bp sequence in the HIV-1 gag p24 region was selected as the target and co-amplified with competitor RNA containing an internal 44-bp deletion. Quantitation of serial dilutions of control RNA samples prepared from the LAI isolate demonstrated a good linearity (R2 = 0.991) within the range between 10 and 250 copies/sample. The detection limit of the assay was determined to be 3.8 copies/sample by Probit analysis and corresponded to 110 copies/ml in plasma. The intra-assay CV value was 9.1%, and the inter-assay value was 25.9%. Both were comparable to those obtained with commercially available HIV-1 RNA quantitation kits. The correlation efficient for the results obtained in 47 plasma samples from HIV-1-infected individuals (subtype A in 1, subtype B in 25, subtype C in 4, subtype F in 1, and CRF01_AE in 16) with the competitive RT-PCR and Cobas Amplicor HIV-1 Monitor test v1.5 was 0.956 for subtype B and 0.947 for subtype non-B. The assay devised is a good alternative for monitoring antiretroviral therapy in resource-poor countries.

Original languageEnglish
Pages (from-to)113-117
Number of pages5
JournalJournal of Virological Methods
Volume142
Issue number1-2
DOIs
Publication statusPublished - 2007 Jun 1

Keywords

  • Competitive RT-PCR
  • HIV-1
  • Monitoring
  • Viral load

ASJC Scopus subject areas

  • Virology

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