Infection time-course and reverse transcriptase assay of HIV/SIV through different coreceptors.

<p>The NP-2/CD4/GPCR cells were seeded and next day inoculated with viruses. Antigen-positive cells were detected by IFA every 3-5 days for up to 7 weeks after inoculation. (<b>A</b>) About 2-3% of the NP-2/CD4/CCR6 cells became antigen positive by IFA after seven days of inoculation of HIV-1HAN2. The ratios of HIV-1-positive cells did not increase after this. Most of the cells became infected conveniently using CCR5 and CXCR4 as coreceptors within seven days. (<b>B</b>) More than 65% cells became HIV-2MIR antigen-positive three weeks after inoculation using CCR6 as cofactor. Major coreceptors took shorter period to reach similar levels of infection (<b>C</b>) NP-2/CD4/CCR6 required seven weeks until most of them became SIVsmE660 antigen-positive. CCR5 took seven days to reach peak levels of infection, whereas CXCR4 did not support infection for SIVsmE660 (<b>D</b>) Culture supernatants of HIV-1HAN2-infected cells were harvested for RT activities at day-7 when only 2.5% of NP-2/CD4/CCR6 cells were IFA-positive. Culture media of NP-2/CD4/CCR5 and NP-2/CD4/CXCR4 cells were measured when 80-90% of them became IFA-positive. NP-2/CD4 cells were examined as negative control. The mean cpm of RT activities for the duplicate samples was calculated by subtracting of mean background cpm value. (<b>E</b>) Spent culture fluids of HIV-2MIR were measured when NP-2/CD4/CCR6 cells reached to 65% antigen-positive, in comparison to about 85% positives of NP-2/CD4/CCR5 and NP-2/CD4/CXCR4 cells (<b>F</b>) RT release of SIVsmE660 was calculated at 80% infection of cells through CCR6 and CCR5 and <0.1% of through CXCR4 as coreceptors.