Anti-HIV Assay and Cytotoxicity Assays. Anti-HIV and
cytotoxicity assays were performed in parallel using the green
fluorescent protein (GFP)-based HOG.R5 reporter cell line that was
constructed and developed specifically for quantitating HIV-1
infectivity. The system was validated and adapted as a moderately
high-throughput procedure for screening natural products for anti-HIV
activity in our laboratory.37,38 Briefly, a reporter cell line for
quantitating HIV-1 replication was developed using HOS (human
osteosarcoma) cells rendered susceptible to HIV-1 infection by the
transfection of genes for CD4 and CCR5, the co-receptor utilized by
macrophage-tropic (R5) HIV-1 isolates. This microtiter assay is based
on the transactivation of a stably integrated HIV-1 LTR-GFP
transcription unit.