We have investigated whether PBMC of HIV-1-seropositive subjects are as susceptible to in vitro infection by HIV-1 as are PBMC from seronegative controls. Accordingly, stimulated PBMC from 19 HIV-1-infected subjects were inoculated with four different variants of HIV-1. None of these cultures produced either detectable quantities of viral reverse transcriptase activity or p24 Ag following inoculation with HIV-1. In contrast, in five of six cases in which these PBMC were depleted of B cells by antibody plus complement prior to viral inoculation, the presence of viral reverse transcriptase and p24 Ag was detected. The presence of normal levels of CD4-Ag at the surface of the CD4+ cells in these populations was established by flow cytometry. Analysis by an immunoblot assay revealed that anti-HIV antibodies were present in the sera obtained from these infected donors; in addition, 7 of 10 culture fluids derived from the nondepleted PBMC were shown to contain virus-neutralizing antibodies. Cultures which were depleted of B cells did not contain detectable levels of antiviral antibodies. Confirmation that the virus produced by the PBMC which had been depleted of B cells was of the strain used to infect the cultures, rather than that which initially caused patient infection, was provided on the basis of differential susceptibility to antibody neutralization. These results suggest that antibodies produced by B cells in cultures of PBMC from seropositive donors may restrict infection by HIV-1 of such cultures under laboratory conditions.