We observed that mixing monoclonal antibodies directed against various epitopes of human chorionic gonadotropin (hCG) can increase the sensitivity of antigen-binding assays. Depending on the antibody pair chosen, the affinity of the mixture was as much as 10-fold higher than either of the monoclonal antibodies assayed separately. Because hCG does not have any repeating sequences, these results do not reflect a change in avidity of an individual antibody. The increased avidity of the mixture can be detected in both solid-phase assays and liquid-phase double antibody radioimmunoassays. This was not a general property of all monoclonal antibodies raised against hCG. Certain antibodies, apparently reacting with the same region of the antigen molecule, do not bind simultaneously and do not result in enhanced affinity when mixed. In addition, certain other pairs of antibodies do not bind cooperatively even though they can bind simultaneously. The mechanism for the increase in affinity depends on the formation of a multicomponent complex. If one of the antibodies of a pair that results in enhanced affinity upon mixing is replaced by its F(ab) fragment, the enhancement is no longer detectable, indicating it is unlikely the enhancement is due to an allosteric effect. Although the F(ab')2 fragment shows some enhancement when mixed with another antibody, it is not as effective as the intact antibody.