Although T lymphocytes are present in the genital mucosa, their function in sexually transmitted diseases is unproven. To determine if cervical T cells mediate HIV-specific cytolysis, mononuclear cells in cytobrush specimens from HIV-1-infected women were stimulated in vitro with antigen. Resultant cell lines lysed autologous targets expressing HIV-1 proteins in 12/19 (63%) subjects, and these responses were detected intermittently on repeated visits. All 8 subjects with blood CD4⁺ counts ⩾500 cells/μl had HIV-1-specific cervical CTL, whereas only 4/11 with counts <500 cells/μl had detectable responses (P = 0.008). Class II MHC– restricted CD4⁺ CTL clones lysed targets expressing Env gp41 or infected with HIV-1. Class I MHC-restricted CD8⁺ clones recognized HIV-1 Gag- or Pol-expressing targets, and the epitopes were mapped to within 9–20 amino acids. Comparisons of intra-individual cervical and blood CTL specificities indicate that epitopes recognized by CTL in the cervix were commonly recognized in the blood. These studies provide the first definitive evidence for an MHC-restricted effector function in human cervical lymphocytes.