Protocol

Diagnosis of Sexually Transmitted Diseases

Volume 903 of the series Methods in Molecular Biology pp 263-271

Date:

Protocol for the Use of a Rapid Real-Time PCR Method for the Detection of HIV-1 Proviral DNA Using Double-Stranded Primer

  • Chou-Pong PauAffiliated withLaboratory Branch, Division of HIV/AIDS Prevention, National Center for HIV/AIDS, Viral Hepatitis, STD, and TB Prevention, Centers for Disease Control and Prevention Email author 
  • , Susan K. WellsAffiliated withLaboratory Branch, Division of HIV/AIDS Prevention, National Center for HIV/AIDS, Viral Hepatitis, STD, and TB Prevention, Centers for Disease Control and Prevention
  • , Timothy C. GranadeAffiliated withLaboratory Branch, Division of HIV/AIDS Prevention, National Center for HIV/AIDS, Viral Hepatitis, STD, and TB Prevention, Centers for Disease Control and Prevention

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Abstract

This chapter describes a real-time PCR method for the detection of HIV-1 proviral DNA in whole blood samples using a novel double-stranded primer system. The assay utilizes a simple commercially available DNA extraction method and a rapid and easy-to-perform real-time PCR protocol to consistently detect a minimum of four copies of HIV-1 group M proviral DNA in as little as 90 min after sample (whole blood) collection. Co-amplification of the human RNase P gene serves as an internal control to monitor the efficiency of both the DNA extraction and amplification. Once the assay is validated properly, it may be suitable as an alternative confirmation test for HIV-1 infections in a variety of HIV testing venues including the mother-to-child transmission testing sites, clinics, and diagnostic testing centers.

Key words

HIV-1 proviral DNA Real-time PCR Double-stranded primer Rapid PCR Nucleic acid testing