An indirect immunofluorescent (IF) test with antigen of partially cleaned Pneumocystis carinii cysts was evaluated for sensitivity and specificity. The test was reactive at the 1:8 and above level for 44% of the persons who had suspected and confirmed infections, for 7.3% of the healthy contacts, and for 1.4% of the general population. Results of tests with sera from people with measurable antibodies to other diseases and those on immunosuppressant therapy suggest that only titers of 1:20 and above are specific. At this level, tests with sera of 32% of the cases, 4% of the contacts, 0% of the healthy controls, and from 0% to 37% (average 5%) of individuals on long-term immunosuppressive therapy or with antibodies to other diseases were positive. The high percentage of positive reactions in cytomegalovirus and in fungal infections could represent double infections. The complement fixation test was less sensitive than the IF test. Antibodies measured by the IF test were immunoglobulin G. No differences in antigenicity were demonstrated by the IF test among the cysts from eight human infections or from induced rat infections.