FIG. 4.

Colocalization of CD63 and HIV Gag in Rab9-depleted cells. (A) Rab9 expression was visualized in parental JC53 cells (left panel) and JC53 Rab9 siRNA cells (right panel) by sequential incubations with a primary anti-Rab9 antibody and an AF488-conjugated secondary antibody. (B) JC53 cells and JC53 Rab9 siRNA cells were transfected with pGag-EGFP and pCD63-mRFP plasmids, and 20 to 60 cells per experiment (n = 3) were optically z-sectioned live at 36 h posttransfection. Cells having <10 or ≥10 intracellular Gag-EGFP puncta colocalizing with CD63-mRFP were classified as having low or high intracellular Gag, respectively. (C) Representative confocal sections of JC53 cells (left panels) and JC53 Rab9 siRNA cells (right panels) cotransfected with pGag-EGFP (green) and pCD63-mRFP (red).