Multiple lines of evidence suggest that cellular immunity plays a critical role in vaccine-induced protection against HIV infection and control of viral replication. Since most new HIV-1 infections are sexually transmitted, the ability to elicit cellular immune responses at mucosal sites is likely to be a crucial property of candidate vaccines. However, there is little information on the ability of candidate AIDS vaccines to induce T cell responses able to home to mucosal sites. Over the past several years, our laboratory has developed several novel methods employing MHC tetramers, ELISPOT and intracellular cytokine staining (ICS) assays for the analysis of SIV-specific CD8+ T cells in gut-associated lymphoid tissues (GALT) and the female reproductive tract. Most recently, we have developed techniques that permit quantitative analysis of CD8+ T cell responses in cervicovaginal biopsies of female rhesus macaques. We now propose to use these new techniques to analyze the ability of two different DNA/MVA vaccination regimens for their ability to induce CD8+ T cell responses that home to genital and gastrointestinal mucosal sites and to protect against vaginal challenge. Specific aims of these studies include: 1. To examine if SIV-specific CD8+ T cells induced by peripheral DNA/MVA immunization home to gastrointestinal and female reproductive tract mucosal sites. 2. To examine if an intravenous MVA boost following DNA priming results in increased homing of CD8+ T cells to genital and gastrointestinal mucosal sites. 3. To examine viral loads and immune responses of DNA/MVA vaccinated animals from Specific Aims # 1 and # 2 after intravaginal challenge with SIVmac251. These experiments should provide valuable information on the ability of candidate AIDS vaccines to induce T cell responses at mucosal sites and improve our knowledge of the signals that mediate homing of T cells to gastrointestinal and reproductive mucosal surfaces.