Characterization of epitopes recognized by cytotoxic T lymphocytes (CTLs) in the sequence of tumor antigens is an important step in the development of tumor therapies. Because carcinoembryonic antigen (CEA) is a protein expressed in a high number of epithelial tumors, it is an interesting target to study. We screened for the presence of HLA-B27-restricted CTL epitopes from CEA by studying the binding to HLA-B27 of 31 synthetic peptides predicted to bind to this molecule. This afforded 16 peptides with moderate or high binding affinity. Immunization of HLA-B27 transgenic mice with the best binder peptides yielded 4 immunogenic peptides: CEA(9-17), CEA(9-18), CEA(138-146) and CEA(360-369). However, splenocytes from mice immunized with a vaccinia virus-expressing CEA recognized only CEA(9-18). These CTLs were of the CD8(+) phenotype, which upon stimulation with peptide specifically produced IFN-gamma. Moreover, they did not cross-react against peptides of region 9-18 from proteins of the CEA family. Our results show that CEA(9-18) may induce specific CTL responses against CEA.