FIG. 12.
(I) Amounts of HIV p24 in supernatants of wild-type (NL4-3) or F12-HIV nef-expressing cells after transfection with the infectious molecular clone HIV-1 NL4-3. A total of 5 × 104 cells were seeded in 24-well plates and, after 24 h, transfected in triplicate with 2 μg of the pNL4-3 plasmid. After an overnight incubation, cell cultures were washed and complete medium was added. Amounts of HIV p24 protein (whose levels 24 h posttransfection were, under all conditions, below the sensitivity threshold of the ELISA) were measured at the indicated times posttransfection (p.t.). (II) Amounts of HIV p24 in supernatants of HeLa CD4 cells cotransfected with the infectious molecular clone HIV-1 pNL4-3 together with vectors expressing wild-type or F12-HIV Nef proteins. A total of 2 × 105 cells were seeded in 6-well plates and after 24 h were cotransfected in duplicate with 1 μg of infectious molecular clone pNL4-3 and a 10-fold molar excess of pcDNAI-based nef-expressing vectors. Measurements of HIV p24 protein were performed at the indicated times posttransfection (p.t.). Values from a representative of three different experiments are reported.