Abstract
Antiviral screens have proved useful for the identification of novel human immunodeficiency virus type 1 (HIV-1) inhibitors. In this study, we describe an HIV-1 full replication (HIV-1 Rep) assay that incorporates all of the targets required for replication in T-cell lines, including the HIV-1 Vif gene. The HIV-1 Rep assay was designed to exhibit optimal sensitivity to late-stage as well as early-stage inhibitors to maximize the likelihood of identification of novel target antiviral compounds in a screen. In addition, the flexibility of the HIV-1 Rep assay allows the rapid evaluation of antiviral compounds against different virus strains in different T-cell lines without significant modification of the assay format. We demonstrate that the HIV-1 Rep assay exhibits characteristics (e.g., a favorable Z' value) compatible with high-throughput screening in a 384-well format. The utility of the HIV-1 Rep assay was demonstrated in a high-throughput screen of >10(6) compounds. To our knowledge, this study represents the first example of an HIV-1 antiviral screen that includes Vif as a functional target and was executed on an industrial scale.