New real-time reverse transcriptase-initiated PCR assay with single-copy sensitivity for human immunodeficiency virus type 1 RNA in plasma
- PMID: 14532178
- PMCID: PMC254331
- DOI: 10.1128/JCM.41.10.4531-4536.2003
New real-time reverse transcriptase-initiated PCR assay with single-copy sensitivity for human immunodeficiency virus type 1 RNA in plasma
Abstract
More sensitive assays for human immunodeficiency virus type 1 (HIV-1) RNA are needed to detect, quantify, and characterize persistent viremia in patients who are receiving antiretroviral therapy and whose plasma HIV-1 RNA levels are suppressed to less than 50 to 75 copies/ml. We therefore developed an internally controlled real-time reverse transcriptase-initiated PCR assay that quantifies HIV-1 RNA concentrations down to 1 copy per ml of plasma. This assay with single-copy sensitivity (the single-copy assay) generates a reproducible linear regression plot of input copy number versus threshold cycle by using HIV-1 RNA transcripts at copy numbers ranging from 1 to 10(6) per reaction mixture. The single-copy assay was compared to the ultrasensitive AMPLICOR HIV-1 MONITOR assay and a more sensitive modification of the ultrasensitive assay by repeatedly testing a low-copy-number panel containing 200 to 0.781 copies of HIV-1 RNA per ml of plasma. This comparison showed that the single-copy assay had a greater sensitivity than the other assays and was the only assay that detected HIV-1 RNA at levels as low as 0.781 copies/ml. Testing of plasma samples from 15 patients who were receiving antiretroviral therapy and who had <75 HIV-1 RNA copies/ml revealed persistent viremia in all 15 patients, with HIV-1 RNA levels ranging from 1 to 32 copies/ml (median, 13 copies/ml). The greater sensitivity of the single-copy assay should allow better characterization of persistent viremia in patients who are receiving antiretroviral therapy and whose HIV-1 RNA levels are suppressed to below the detection limits of present assays.
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References
-
- Coffin, J. M. 1995. HIV population dynamics in vivo: implications for genetic variation, pathogenesis, and therapy. Science 267:483-489. - PubMed
-
- Collins, M. L., B. Irvine, D. Tyner, E. Fine, C. Zayati, C. Chang, T. Horn, D. Ahle, J. Detmer, L. P. Shen, J. Kolberg, S. Bushnell, M. S. Urdea, and D. D. Ho. 1997. A branched DNA signal amplification assay for quantification of nucleic acid targets below 100 molecules/ml. Nucleic Acids Res. 25:2979-2984. - PMC - PubMed
-
- Dornadula, G., H. Zhang, B. VanUitert, J. Stern, L. Livornese, Jr., M. J. Ingerman, J. Witek, R. J. Kedanis, J. Natkin, J. DeSimone, and R. J. Pomerantz. 1999. Residual HIV-1 RNA in blood plasma of patients taking suppressive highly active antiretroviral therapy. JAMA 282:1627-1632. - PubMed
-
- Elbeik, T., E. Charlebois, P. Nassos, J. Kahn, F. M. Hecht, D. Yajko, V. Ng, and K. Hadley. 2000. Quantitative and cost comparison of ultrasensitive human immunodeficiency virus type 1 RNA viral load assays: Bayer bDNA quantiplex versions 3.0 and 2.0 and Roche PCR Amplicor monitor version 1.5. J. Clin. Microbiol. 38:1113-1120. - PMC - PubMed
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