The lifespan and survival of dendritic cells (DC) in vivo are potentially critical to the expansion of T cell immune responses. We have previously reported that DC loaded with specific antigen are rapidly eliminated by cytotoxic T lymphocytes (CTL) in vivo, but the site, mechanism, and consequences of DC elimination were not defined. In this article we show that DC elimination in vivo occurs in a perforin-dependent manner and does not require IFN-gamma or the presence of CD4(+)CD25(+) regulatory T cells. Most importantly, failure to eliminate DC had profound consequences on the CTL immune response. Perforin-deficient mice showed a progressive increase in the numbers of antigen-specific CD8(+) T cells after repeated immunizations with DC. In contrast, in control mice the number of antigen-specific CD8(+) T cells did not notably increase with repeated immunizations. Lastly, we also show that CTL-mediated elimination of DC occurs in peripheral tissues but not in the lymph node. Our data suggest that CTL act as "gatekeepers" that control access of antigen-loaded DC into the lymph node, thereby preventing continued expansion of antigen-specific T cells.