Murine monoclonal antibodies to flagellar, surface membrane and cytoplasmic antigens of New World Leishmania were assessed for their taxonomic specificity in enzyme-linked immunosorbent assays with three genera of the family Trypanosomatidae and three species and seven subspecies of the genus Leishmania. Antibodies exhibiting exclusive reactivity with either the flagellum, flagellar pocket, kinetoplast, or nucleus lacked specificity at all phylogenetic levels and, in fact, recognized epitopes common to cultured mammalian cells. Monoclonals to intracellular antigens were capable of distinguishing Leishmania from Trypanosoma and Endotrypanum. Antibodies reactive at the surface membrane could separate six isolates of L. braziliensis from three isolates of L. mexicana but the differences in antigen expression were frequently quantitative rather than qualitative. Antigenic variability within species and/or subspecies often exceeded that which was observed between species and/or subspecies. At least one monoclonal antibody was specific for a surface antigen peculiar to a subpopulation of promastigotes of an L. braziliensis panamensis isolate.