Determination of HIV infectivity in vitro and its inhibition by antiretroviral drugs by monitoring reduction of production of p24 antigen is expensive and time consuming. Such assays also do not allow accurate quantitation of the number of infected cells over time. To develop a simple, rapid, and direct method for monitoring HIV infection, we generated a stable T-cell line (CEM) containing a plasmid encoding the green f luorescent protein (humanized S65T GFP) driven by the HIV-1 long terminal repeat. Clones were selected that displayed low constitutive background f luorescence, but a high level of GFP expression upon infection with HIV. HIV-1 infection induced a 100-to 1,000-fold increase in relative f luorescence of cells over 2 to 4 days as monitored by f luo-rescence microscopy, cytof luorimetry, and f low cytometry. Addition of inhibitors of reverse transcriptase, protease, and other targets at different multiplicities of infection permitted the accurate determination of drug susceptibility. This tech-nique also permitted quantitation of infectivity of viral prep-arations by assessment of number of cells infected in the first round of infection. In conclusion, the CEM-GFP reporter cell line provides a simple, rapid, and direct method for monitor-ing HIV infectivity titers and antiretroviral drug susceptibility of syncytium-inducing strains.
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