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HIV inhibition assay. A serial diluted drug (10 μM, 1 μM, 100 nM, 10 nM, 1 nM, and 0.1 nM), was tested against 100 TCID50 HIVADA infection in a pseudovirus-based assay. The assay quantifies the activity of a drug to inhibit HIV-induced reporter luciferase activity. The viral infection was determined on day 3 by measuring the reporter luciferase activity in TZM. CD4+. CCR5+ cells post-infection using commercially available kits. Antiviral data were reported as the quantity of drug required to inhibit production by percentage. Controls include (1) infected cells without adding any drugs as a baseline, (2) pseudovirus with non-specific envelope (e.g., HCV) which cannot infect TZM. CD4+. CCR5+ cells, and (3) an HIV-specific RT inhibitor as a positive control (e.g. AZT). The cell toxicity of the compounds against HIV was evaluated by similar microtiter assays with TZM. CD4+. CCR5+ and Vero cells. Serial diluted drugs (10 mM, 1 mM, 100 μM, 10 μM, 1 μM and 0.1 μM) were tested in cells using the cell viability assay which is commercially available (Promega). The toxicity data are reported as the quantity of a drug required to reduce cell viability by percentage.
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