Both IgG and secretory IgA Abs in mucosal secretions have been implicated in blocking the earliest events in HIV-1 transit across epithelial barriers, although the mechanisms by which this occurs remain largely unknown. In this study, we report the production and characterization of a human rIgA(2) mAb that carries the V regions of IgG1 b12, a potent and broadly neutralizing anti-gp120 Ab which has been shown to protect macaques against vaginal simian/HIV challenge. Monomeric, dimeric, polymeric, and secretory IgA(2) derivatives of b12 reacted with gp120 and neutralized CCR5- and CXCR4-tropic strains of HIV-1 in vitro. With respect to the protective effects of these Abs at mucosal surfaces, we demonstrated that IgG1 b12 and IgA(2) b12 inhibited the transfer of cell-free HIV-1 from ME-180 cells, a human cervical epithelial cell line, as well as Caco-2 cells, a human colonic epithelial cell line, to human PBMCs. Inhibition of viral transfer was due to the ability of b12 to block both viral attachment to and uptake by epithelial cells. These data demonstrate that IgG and IgA MAbs directed against a highly conserved epitope on gp120 can interfere with the earliest steps in HIV-1 transmission across mucosal surfaces, and reveal a possible mechanism by which b12 protects the vaginal mucosal against viral challenge in vivo.