FIG 8.

BN-PAGE gel mobility shift of Comb-mut and CM-LK trimeric Env in the presence of various bnAbs. (A) Comb-mut and CM-LK virions, displaying >95% cleaved Env, were incubated in the presence of the IgGs specified (25 μg/ml) and analyzed using BN-PAGE Western blot. Binding of an antibody causes the Env trimer to run more slowly on the gel, thus shifting the band upward on the blot. 2G12 and DEN3 were used as positive- and negative-control antibodies, respectively. (B) Virion-antibody complexes were fixed using the chemical cross-linker BS³—after antibody binding but before adding detergent—to prevent antibodies from falling off during the gel run. (C) Comb-mut and CM-LK virions were incubated with 447-52D or F425-B4e8 IgGs that bind to V3, or control IgG 2G12, and then Env trimers were solubilized using detergent and analyzed using a BN-PAGE Western blot.