Human immunodeficiency virus type 1 (HIV-1) infection elicits neutralizing antibodies directed against two discrete regions of the gp120 exterior envelope glycoprotein: the third variable (V3) loop and the CD4 binding region. Monoclonal antibodies directed against these two regions demonstrated additive or, in some cases, weakly synergistic neutralization of HIV-1 infection. Cooperativity in virus neutralization was also observed for some gp120 mutants that, in the absence of anti-V3 loop antibodies, were relatively resistant to neutralization by antibodies directed against the CD4 binding region. Although the binding of some anti-V3 region monoclonal antibodies increased the recognition of the multimeric envelope glycoproteins by anti-CD4 binding antibodies, this enhanced binding was not predictive of the degree of cooperativity observed in virus neutralization. These results suggest that elicitation of both types of neutralizing antibodies should increase the efficacy of vaccine preparations.