Intravenous inoculation of irradiated virulent promastigotes of Leishmania mexicana, or intradermal inoculation of avirulent temperature-sensitive clones of the same parasite, can protect BALB/c mice from progressive disease when challenged with parental organisms. However, if animals are prechallenged s.c. with irradiated parental parasites before (or shortly after, i.e., within 10 days) any other immunization regime is used, the s.c. challenge effectively suppresses development of protective immunity. The role of N-linked sugars on promastigotes in providing the determinants responsible for protection and/or suppression of protection in these assays has been examined. Growth of parasite in 2 micrograms/ml tunicamycin has no effect on incorporation of [3H]leucine but decreases incorporation of [3H]mannose by some 40 to 50%. Such tunicamycin-treated avirulent clones or irradiated virulent organisms are now unable to induce a protective response. However, tunicamycin-treated parental parasites given s.c. could still suppress the protection seen when irradiated parasites were given i.v. as immunogen. Tunicamycin-treated avirulent organisms given s.c. subcutaneously unlike the untreated avirulent clones, now also caused suppression of protection. These data suggest that the determinants responsible for development of protective immunity to L. mexicana in BALB/c mice are dependent on N-linked glycoproteins for their expression, unlike the determinants responsible for suppression of that protection. By using swainsonine as an alternative inhibitor of N-linked glycoprotein synthesis, the data additionally suggest that no complex processing of N-linked sugars takes place to reveal the immunogenic determinants in this system.