Externally oriented surface membrane constituents of promastigotes from several Leishmania species were radiolabeled with 125I. Autoradiographs of cell surface-labeled and sodium dodecyl sulfate-polyacrylamide gel electrophoresis-separated proteins of the stocks revealed distinctive patterns of bands in the molecular weight range of 6,000 to 240,000. Immunoprecipitation of detergent extracts of the labeled promastigote stocks with anti-Leishmania donovani membrane serum demonstrated that each of the stocks contained some antigenically cross-reactive determinants. The electrophoretic patterns of these determinants serve both to distinguish the parasite stocks (by unique, species-specific patterns) and to indicate antigenic similarities in stocks thought to be different by other biochemical criteria. At least 12 cross-reactive cell surface antigens in two New World leishmanias are recognized by polyvalent anti-L. donovani serum, suggesting that these common leishmanial antigens may account for the documented serological cross-reactivities among various Leishmania species. In all stocks tested, an iodinated protein was identified which had a relative molecular weight of 65,000 under reducing conditions but which demonstrated an increase in relative mobility in sodium dodecyl sulfate-polyacrylamide gels under nonreducing conditions. Distinctive patterns of the antigens common to the several stocks were also demonstrated with the use of monoclonal antibodies.