Seminal HIV-1-specific cytotoxic T lymphocytes (CTLs) could provide an important immune defense against local HIV-1 infection, and be important in impeding the spread of HIV-1 infection. In this study, we demonstrate that autologous blood-derived dendritic cells (DCs) loaded in vitro with synthetic HIV-1 peptides representing known CTL epitopes activated HLA class I restricted, anti-HIV-1 CTLs and interferon gamma responses in seminal CD8+ T cells from subjects with chronic HIV-1 infection on antiretroviral therapy. CTLs specific for the same HLA-restricted epitopes were detected in semen and blood of the same individuals by stimulation with peptide-loaded DCs. Anti-HIV-1 CTL responses from semen were enhanced by stimulation with DCs loaded with HIV-1 peptides and interleukin 12. Our results suggest that blood-derived DCs have HIV-1 antigen-presenting capacity for seminal CTL in HIV-1-infected subjects. The DC-T cell system can serve as a model for immunotherapy of HIV-1 infection in the local genital tract as well as systemic blood circulation.