Figure 6

Cytofluorimetric analysis of VLPs released by 18-4s cells. (A) Hundred microliters of supernatants from induced 18-4s cells were clarified, adjusted to 400 ng of CAp24 contents, two-fold serially diluted in complete medium, incubated with aldehyde latex beads, and finally analyzed by FACs (slopes b). As control, supernatants from 293/RGP cells adjusted for equal VLP contents were used (slopes a). (B) The supernatants from induced 18-4s (slope b) and, as control, from 293/RGP cells (slope a) were cleared from a total of 400 ng of VLPs by ultracentrifugation, and then analyzed by FACs upon incubation with aldehyde latex beads. (C) Hundred ng of 18-4s VLPs were incubated with aldehyde latex beads and labeled with either KC57 PE-conjugated anti-CAp24 (on the left) or anti-Nef mAbs # 6.2 (on the right) and, in a second step, with PE-conjugated goat anti-mouse IgG. Histograms refer to the FACs analysis of aldehyde latex beads incubated with supernatants of 18-4s cells either non induced (a) or induced but in the absence (b) or presence (c) of the permeabilization step before the Ab labeling.