Diagnosis of Sexually Transmitted Diseases pp 263-271

Part of the Methods in Molecular Biology book series (MIMB, volume 903)

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 Pau
  • Susan K. Wells
  • Timothy C. Granade
Protocol

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 

References

  1. 1.
    Stekler JD, Swenson PD, Coombs RW et al (2009) HIV testing in a high-incidence population: is antibody testing alone good enough? Clin Infect Dis 49:444–453PubMedCrossRefGoogle Scholar
  2. 2.
    Ou CY, Yang H, Balinandi S et al (2007) Identification of HIV-1 infected infants and young children using real-time RT PCR and dried blood spots from Uganda and Cameroon. J Virol Methods 144:109–114PubMedCrossRefGoogle Scholar
  3. 3.
    Morris SR, Little SJ, Cunningham T, Garfein RS, Richman DD, Smith DM (2010) Evaluation of an HIV nucleic acid testing program with automated Internet and voicemail systems to deliver results. Ann Intern Med 152:778–785PubMedGoogle Scholar
  4. 4.
    El Ekiaby M, Lelie N, Allain JP (2010) Nucleic acid testing (NAT) in high prevalence-low resource settings. Biologicals 38:59–64PubMedCrossRefGoogle Scholar
  5. 5.
    Stevens WS, Marshall TM (2010) Challenges in implementing HIV load testing in South Africa. J Infect Dis 201(Suppl 1):S78–S84PubMedCrossRefGoogle Scholar
  6. 6.
    Pau CP, Wells SK, Rudolph DL, Owen SM, Granade TC (2010) A rapid real-time PCR assay for the detection of HIV-1 proviral DNA using double-stranded primer. J Virol Methods 164:55–62PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2012

Authors and Affiliations

  • Chou-Pong Pau
    • 1
  • Susan K. Wells
    • 1
  • Timothy C. Granade
    • 1
  1. 1.Laboratory Branch, Division of HIV/AIDS PreventionNational Center for HIV/AIDS, Viral Hepatitis, STD, and TB Prevention, Centers for Disease Control and PreventionAtlantaUSA

Personalised recommendations