The Ca²⁺-dependent protease antisera and the purified specific antibodies from Allomyces arbuscula have shown very specific recognition when blotted against the total protein extract or the purified 43-40 kDa Ca²⁺-dependent protease from this aquatic fungus. By immunoblotting and immunofluorescence techniques using specific antibodies, we have shown that the enzyme activity is developmentally regulated and is related to the presence of antigen and not to any specific inhibitor. The immunofluorescence was absent in zoospores but appeared in polarized forms in germinating spores. In elongating hyphae the protease was mainly localized along the cytoplasmic membrane and in the cytoplasm, with predominance at the apex.