TAS contributes to the exposure of binding sites within gp41 necessary for inhibition by C34 peptide. MAGI +/+ cells were plated at 10,000 cells/well, and coincubated with HIV-1NL4.3 at 4°C for 2 hours. Unbound virus was washed with PBS, and cells were shifted to 23°C-TAS for 3 hours. Fusion inhibitors sCD4 (PRO 542) and C34 peptide were either added at the onset of 23°C incubation (white bars) or after TAS had been established for 2 hours (black bars). Inhibitors were allowed 1 hour for binding, then washed. Following the 3 hour TAS incubation, the cells were washed with PBS and shifted to 37°C. The averages of triplicate experiments are shown (n = 4). Error bars represent the standard error of the mean. Relative infectivity was quantified using a liquid β-Gal assay. OD 405 nm, optical density at 405 nm. The concentrations of the inhibitors were: sCD4 (5 ug/mL) and C34 (100 nM).