FIG. 4.

Membrane flotation analysis of RT proteins. 293T cells were transfected with pGAG, GPfsPR⁻, or the designated expression constructs. Two days posttransfection, cells were harvested, homogenized, and subjected to equilibrium flotation centrifugation analysis as described in Materials and Methods. Ten fractions were collected from the top downward. Fraction aliquots were resolved by SDS-PAGE (10%) and probed with a monoclonal antibody directed against HIV-1 p24CA or the HA tag. During ultracentrifugation, membrane-bound Gag proteins floated to the 10 to 65% sucrose interface and became enriched in fraction 3.