The biological basis for the specificity of host infectivity patterns of Cryptosporidium spp., in particular C. hominis and C. parvum, has yet to be fully elucidated. Comparison of the C. parvum and C. hominis P23 and GP900 predicted amino acid sequences revealed 3 differences in P23 and 4 and 17 differences in GP900 domains 1 and 5, respectively. Using monoclonal antibodies developed against the surface (glyco)proteins P23 and GP900 of the C. parvum Iowa isolate, solubilized glycoprotein from three C. hominis isolates was screened for reactivity using Western immunoblots. One of ten P23 MAbs and three of 21 GP900 MAbs were not reactive with any of the three C. hominis isolates. The non-reactive P23 MAb binds to a peptide epitope, while the non-reactive GP900 MAbs bind to either carbohydrate/carbohydrate-dependent or peptide epitopes of C. parvum. These results demonstrate phenotypic differences between C. hominis and C. parvum within two (glyco)proteins that are involved in parasite gliding motility and attachment/invasion.