Tat is released by HIV-1–infected cells and binds the cell surface of producing and nearby cells. (A) PBMCs were acutely infected with either III B or Ba-L HIV-1 strains, and Tat amounts (♦) were evaluated both in culture supernatants (by ELISA) and on the cell surfaces (by immunostaining and cytofluorimetric analysis). Numbers represent percent of Tat-positive cells at the indicated time points. The infection was followed for 17 days and monitored as RT activity in the cell supernatants (□). An experiment is shown as representative of 3 performed with comparable results. (B) Anti-Tat NT3 2D1 surface staining of U937/Tat cells and of C8166 cells after coculture with U937/Tat cells. U937/PINCO and C8166 cells are shown as negative controls. These stainings are representative of 3 experiments with similar results. (C) Cytofluorimetric evaluation of rTat binding to the surface of C8166 cells, shown as percent of Tat-positive cells. (D) Identification of Tat region(s) involved in binding the cell surface. C8166 cells were incubated with the described GST-Tat variants, and the amount of Tat-positive cells was detected by cytofluorimetric analysis using an anti-GST mAb. (*P ≤ .01 versus binding of GST-Tat₈₆). In panels C and D, each value indicates mean ± SD of 2 experiments performed in triplicate.