HMB45 is a melanoma-specific monoclonal antibody that has found widespread use in diagnostic pathology. Recent reports, however, have suggested that this antibody may cross-react with a small number of carcinomas and other epithelial cells. The authors tested the hypothesis that these latter reports represent examples of false-positive immunostaining by comparing the immunostaining on breast, salivary gland, and lung tumors with the following: (1) a commercial ascites preparation of this monoclonal antibody; (2) a protein A-purified antibody preparation derived from ascites fluid; and (3) supernatant fluid obtained from the hybridoma cell line. The authors found that all examples of nonmelanoma immunostaining in the carcinomas tested were eliminated with the nonascites fluid preparations, whereas strong immunostaining of melanomas was retained. The authors conclude that contaminated commercial ascites fluid preparations of HMB45 may account for most, if not all, of the reports of nonmelanoma immunostaining with HMB45.