ABSTRACT.Immunological relationships between genera and species of microsporidia were examined by immunoblot analysis. Exospore polypeptides from twoNosemaspp., threeVairimorphaspp., and two undescribedVairimorpha‐like isolates were analyzed. Gel electrophoresis and immunoblot analysis revealed that a variety of polypeptides, mostly between 15,000 and 90,000 molecular weight, are present on the exospore. The threeVairimorphaspp. are closely related immunologically to each other, but less so to the two undescribedVairimorpha‐like isolates. The twoNosemaspp. are immunologically distant from each other and from theVairimorphaspp. Indirect evidence, however, indicated that many internal spore polypeptides present in both genera are similar. Cross‐reactivity between exospore polypeptides from entomophilic microsporidia and antisera to a mammalian microsporidium,Encephalitozoon cuniculi, was very limited. These results indicate that immunoblot analysis of exospore polypeptides may be employed to investigate the interrelatedness of microsporidian species, and that exospore polypeptides of some microsporidia are sufficiently diverse to be of immunodiagnostic