ObjectivesWe examined 96 bronchoalveolar lavage fluid (BALF) specimens from AIDS patients with provenPneumocystis cariniipneumonia (PCP) in order to compare the relationship of organism burden, viability and antigen expression with disease severity at the time of clinical presentation.MethodsTinctorial analysis of BALF specimens with proven PCP using Diff-Quik, cresyl echt violet and erythrosin B stains to evaluate organism burden and viability.P. cariniiantigen examination was performed by Western blot analysis.ResultsP. cariniicluster ratios were more sensitive than cyst counts as an indicator of organism burden, and correlated well with the alveolar-arterial oxygen gradient as a measure of disease severity. Erythrosin B, the vital stain used to measureP. cariniiviability, displayed a wide range of values and provided little useful information. Antigens of 35–45 and 95 kD, which were specific forP. carinii, were found by immunoblot analysis in BALF cellular fraction of most patients with pneumocystosis. By contrast, antigens of 52 and 66 kD, which were found in both BALF supernatant and cellular fractions ofP. cariniipatients and controls, most likely represented albumin and immunoglobulin G heavy chain, respectively, of host origin. The 35–45 kD antigen was found in 88% of the BALF specimens and appeared to represent an important marker ofP. cariniiinfection. The 95 kD antigen was detected in 49% of the specimens.ConclusionsWe conclude that analysis ofP. cariniicharacteristics in BALF specimens of patients with pneumocystis may provide additional information. These data will also be helpful in developing more sensitive assays and in targeting specificP. cariniifactors for future investigation.