Comparison of methods to quantify inducible HIV-1 outgrowth.

The quantitative viral outgrowth assay (qVOA) is the gold standard for measuring inducible, replication-competent HIV-1. Using MOLT4-R5 and SupT1-R5 cell lines instead of allogeneic blasts and HIV-1 RNA detection rather than p24 enzyme-immunoassay (EIA) has been proposed to improve the sensitivity of the qVOA. It is unclear, however, how these alternative approaches affect qVOA performance. We compared three qVOAs methods across 15 persons with HIV-1 on suppressive antiretroviral therapy and found that the MOLT4-R5 method yielded a significantly higher proportion of p24-positive wells (42%) than both the allogeneic blast (29%) and SupT1-R5 (32%) assays. Additionally, 5 of 7 qVOAs that were negative by p24 EIA showed viral outgrowth by HIV-1 RNA quantification (>10-fold increase within 7 days). These findings reveal the potential for underestimation of the latent, inducible reservoir by qVOA depending on the target cells used and the measure of viral outgrowth. Use of MOLT4-R5 cells with both p24 EIA and HIV-1 RNA to detect viral outgrowth was the most sensitive method.