In this report we describe the first purification and characterization of the acid alpha-mannosidase from the human parasite Trypanosoma cruzi. The purified enzyme exhibited a native mol. wt of 240,000 Da and is apparently composed of four identical subunits of mol. wt 58,000 Da. Each of the four subunits contains one N-linked high-mannose-type oligosaccharide. The alpha-mannosidase exhibited a pH optimum of 3.5 and a pI of 5.9. This low pH optimum and the ability of swainsonine to inhibit its activity suggest that the alpha-mannosidase is a lysosomal enzyme. Antibodies against the T.cruzi enzyme did not react with mammalian lysosomal alpha-mannosidase and, conversely, antibody against a rat lysosomal alpha-mannosidase did not react with the T.cruzi enzyme. Thus, the T.cruzi enzyme appears to be distinct from its mammalian counterpart.