Supporting Figure 5
Fig. 5. BMS-806 does not inhibit envelope glycoprotein CD4 binding. We transfected 293T cells with pSVIIIenv plasmids expressing either JR-FL or YU2 soluble gp120 envelope glycoproteins. Seventy-two hours later, the supernatant was harvested and used for the ELISA assay. The ELISA was performed using 2% milk proteins/10% FBS in PBS instead of SuperBlock. Antibodies or (His)6-tagged soluble CD4 (sCD4) were incubated with the captured gp120 in the absence or presence of different concentrations of BMS-806 for 1 h. The plates were then washed, and bound antibody was detected by using peroxidase-conjugated goat anti-human IgG (Ig-constant region-specific) antibody (Sigma). In the case of sCD4 binding, the plates were washed and incubated with penta-His antibody (Qiagen) for 1 h, followed by peroxidase-conjugated goat anti-mouse IgG (Sigma). The values shown represent the means and SEs of duplicate points in one assay. The results are typical of those obtained in two independent experiments.
