The taxonomic status of Pneumocystis carinii is uncertain, and P. carinii has been categorized both as a fungus and as a protozoan. Recent comparisons of RNA sequence homologies between P. carinii and several genera of fungi and protozoa suggest that P. carinii has closer affinities with the ascomycetes than with the protozoa. The translatory systems of the fungi, however, require three soluble protein factors for peptide chain elongation rather than the two necessary in other eukaryotic systems; to date the additional protein elongation factor (EF-3) appears to be unique to fungi. Western blot analysis of cell-free extracts of P. carinii, derived from rat, was done using a polyclonal antibody raised in rabbits to Saccharomyces cerevisiae EF-3. Anti-EF-3 cross-reacting material was detected only in lysates of Candida albicans and S. cerevisiae included as fungal controls; no cross reaction was detected in lysates of P. carinii, P. carinii-infected rat lung, or a protozoan control (Trichomonas vaginalis).