QUANTIFICATION OF HIV-1 PROVIRAL DNA BY A STANDARDIZED COLORIMETRIC PCR-BASED ASSAY

A simple method was developed for measuring human immunodeficiency vir us type 1 (HIV-1) proviral DNA in mononuclear cells based on the comme rcially available Amplicor(TM) HIV-1 polymerase chain reaction (PCR) a ssay and the limiting dilution method. The lowest limit of detection w as four proviral genomes per 10(6) cells. The accuracy was demonstrate d by using serial dilutions of LAV-8E5 cells, and the interassay varia bility was 0.2 log. The technique was used to measure HIV-1 proviral D NA in the peripheral blood mononuclear cells (PBMC) of 18 antiretrovir al drug-naive HIV-1-positive individuals before and 4 weeks after init iating double nucleoside therapy. The DNA proviral titers at baseline (median = 3.45, range = 2.11-4.7 log copies/10(6) cells) were 2.08 log greater than the infectivity titers, but there was a correlation betw een these two parameters (r = 0.63, P = 0.009). The mean decrease in t he proviral DNA titer after 4 weeks of therapy was 0.31 log, whereas t he decrease in the infectivity titer was 0.81 log and the decrease in the plasma RNA concentration was 1.29 log. The technique was also used to measure HIV-1 proviral DNA in the PBMC of 11 patients who had unde tectable plasma HIV-1 RNA after being placed on combination antiretrov iral therapy. Although proviral DNA remained detectable in all patient s after 36 weeks of treatment, a gradual decline with an estimated hal f-life of 21-58 weeks was observed. The reliability of this simple and convenient colorimetric PCR-based technique indicates its suitability for assessing the effect of current antiretroviral regimens on the la tent reservoirs of provirus. (C) 1998 Wiley-Liss, Inc.