The diagnosis of Histoplasma capsulatum infection by serologic testing for the presence of antibodies is limited by a high rate of false positive and false negative results and by the requirement that the patient have a normal immune response. We have developed a radioimmunoassay for the detection of H. capsulatum antigen in urine and serum specimens. Antigenuria was noted in 20 of 22 episodes of disseminated histoplasmosis that occurred in 16 patients, in 6 of 32 patients with self-limited infection, in 2 of 32 patients with cavitary histoplasmosis, and in 4 of 8 patients with a sarcoid-like illness caused by H. capsulatum. The detection of antigen in urine was reproducible in 38 of 41 (93 percent) retests of specimens. H. capsulatum antigen was also detected in the serum during 11 of the 22 episodes of disseminated histoplasmosis, in none of the 12 episodes of other types of histoplasmosis in patients with antigenuria, in 1 of the 33 patients with histoplasmosis who lacked the urinary antigen, and in none of the 50 controls. Antigenemia and antigenuria decreased after initiation of antifungal therapy and recurred in patients who had a relapse. We conclude that this radioimmunoassay for H. capsulatum antigen represents a useful new method for the rapid diagnosis of disseminated histoplasmosis.