Both a conserved surface metalloprotease of leishmania, gp63 as well as certain gp63-derived peptides, have been shown to have immunoprophylactic potential in mouse models of leishmaniasis. In addition, PBMC from individuals with cutaneous, mucosal, or cured visceral leishmaniasis respond in vitro to both native and rgp63. In this report, we mapped human T cell epitopes within gp63. T cells from leishmaniasis patients responded in vitro to certain peptides of gp63 by proliferation and IFN-gamma production. One peptide, (PT7), stimulated cells from all individuals tested (n = 7). Anti-PT7 T cell lines derived from PBMC of a mucosal leishmaniasis patient contained a heterogeneous population of cells which responded by proliferation and IFN-gamma production to in vitro stimulation with Leishmania promastigote lysate. Another peptide (PT1) derived from Leishmania chagasi gp63 stimulated PBMC from an L. chagasi patient although the corresponding Leishmania major-derived peptide did not. Both L. major PT7 and L. chagasi PT1 were able to induce anti-Leishmania-specific T cell lines from normal human PBMC. These T cell lines responded to in vitro stimulation with promastigote lysate indicating that both peptides were immunogenic for naive T cells in vitro. In conclusion, both antigenic and immunogenic gp63 peptide sequences have been defined, some appearing to be conserved among Leishmania species and at least one that appears to be species specific.