The specificity and functional activity of antibodies to human immunodeficiency virus (HIV) in the sera of MRL/lpr mice were analyzed by peptide-ELISA, antibody-dependent cellular cytotoxicity, and virus neutralization. The specificity of the antibodies in the sera of autoimmune MRL/lpr mice was similar to that of HIV-infected persons; the mouse sera specifically recognized the V3 loop of gp120 and immunodominant regions of gp41 and p24. Moreover, such binding was inhibited both by human HIV-positive sera and by soluble peptides in competition experiments. MRL/lpr sera displayed anti-HIV antibody-dependent cellular cytotoxicity using human peripheral blood lymphocytes as effector cells and HIV-infected H9 cells. Furthermore, the fact that MRL/lpr sera neutralized in vitro infectious HIV (both strains IIIB and MN) suggests these antibodies recognize viral epitopes on the membrane of infected T cells.