FIG 8.
bNAb-mediated Env internalization is dynamin dependent. Cell surface staining of primary CD4+ T cells infected with either NL4.3 GFP ADA-based virus (A) or CH58 T/F virus (B) was performed with A32, 17b, PGT151, PGT126, PG9, and 2G12 MAbs in the presence or absence of 50 μM DIP. (Left) Quantification of remaining antibody-Env complexes on the cell surface over different time points (90 and 180 min) is expressed as a percentage of MFI relative to the 0-min time control. (Right) AUC values were calculated based on MFI data sets using GraphPad Prism software. Cell surface staining of 293T cells transfected with HIV-1JRFL Env, along with the lamin B receptor-CFP plasmid, was performed to locate the nuclear envelope with Alexa Fluor 594-conjugated 2G12 MAb in the presence or absence of 40 μM DIP. (C) Images show the localization of antibody-Env complexes at different time points (0, 60, 120, and 180 min). Images represent a single confocal z-section through the middle of the cell; 20 cells were imaged per condition, and representative images are shown. Scale bar, 10 μm. (D) Quantification of remaining antibody-Env complexes over different time points is expressed as the percentage of surface fluorescence relative to the 0-min control. Error bars indicate means ± the SEM. Statistical significance was tested using an unpaired t test or a Mann-Whitney U test based on statistical normality (*, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001).