ABSTRACT.Total RNA fromPneumocystis cariniiobtained directly from the rat lung and from short term culture on A549 cells was evaluated for size and purity. An isolation procedure using guanidine isothiocyanate and lithium chloride was preferable to a hot phenol method. Host cells were eliminated by hypotonic lysis and a series of microfiltrations.Pneumocystis cariniiwere pretreated with Zymolyase for increased susceptibility to chaotropic agents. The major ribosomal species ofP. cariniiRNA migrated similarly toSaccharomyces cerevisiaerRNA. The 28s‐like species migrated well ahead of rat and A549 cell rRNA and well behind the prokaryotic large rRNA specie