Persistent infection with oncogenic types of human papillomaviruses (HPV) is the major cause of cervical cancer precursor lesions. Cellular immune responses are considered important in the elimination of HPV infection, but the targets are not well defined. HPV E1 and E2 proteins form a replicative complex necessary for viral genome maintenance. To investigate whether epitopes in the E1 or E2 proteins can serve as targets for cytotoxic T-lymphocyte (CTL)-mediated killing, we identified peptides containing the human leukocyte antigen (HLA)-A*0201 binding motif in the deduced amino acid sequences of the HPV-16 E1 and E2 genes. Binding affinity of the peptides was measured by HLA-A*0201 up-regulation on T2 cells. Peptides with high binding-affinity were tested for their ability to elicit peptide-specific CTLs from healthy blood donors. We found one peptide from the E1 and one from the E2 protein sequence that were capable of eliciting peptide-specific CTLs. The E2-specific CTLs lysed an HPV-16-transfected cervical carcinoma cell line, but not the untransfected HPV-negative parental cell line, indicating that the identified E2 epitope can be presented to CTLs in HPV-positive epithelial cells. These findings might have potentially important implications for studies of the natural history of HPV infection in relation to cervical carcinogenesis.