Human immunodeficiency virus type 1 (HIV-1) envelope glycoproteins (gp120 and gp41) elicit virus-neutralizing antibodies (VNAB) and also antibodies enhancing HIV-1 infection (EAB). Several epitopes eliciting VNAB have been defined, the principal virus-neutralizing determinant being assigned to the V3 loop of gp120. To provide a background for a rational design of anti-HIV vaccines, it also appears important to define domains eliciting EAB. This was accomplished by screening antisera against synthetic peptides covering almost the entire sequence of gp120/gp41 for their enhancing effects on HIV-1 infection of MT-2 cells, a continuous T cell line. Many (16/30) of the antisera significantly enhanced HIV-1 in the presence of human complement. Antibodies to complement receptor type 2 (CR2) abrogated the antibody-mediated enhancement of HIV-1 infection. Antisera to V3 hypervariable loops of 21 distinct HIV-1 isolates were also tested for their enhancing effects on HIV-1IIIB infection. 11 of these sera contained VNAB and 10 enhanced HIV-1IIIB infection. All antisera with virus-enhancing activity contained antibodies crossreactive with the V3 loop of HIV-1IIIB, and the virus-enhancing activity increased with increasing serological crossreactivity. These results suggest that immunization with antigens encompassing V3 loops may elicit EAB rather than protective antibodies if epitopes on the immunogen and the predominant HIV-1 isolate infecting a population are insufficiently matched, i.e., crossreactive serologically but not at the level of virus neutralization.