LPS is a natural adjuvant that potentiates Ag-specific T cell survival and Th1 differentiation by stimulating MyD88 and Toll/IL-1R domain-containing adaptor-inducing IFN-beta (TRIF) signaling pathways. In this study, we reveal the TRIF pathway is critical for amplifying murine effector T cell accumulation into nonlymphoid tissues following immunization with Ag plus LPS. Although LPS increased the accumulation of splenic T cells in TRIF-deficient mice, markedly fewer T cells were recovered from liver and lung in comparison to wild type. Most of the T cells primed in TRIF-deficient mice failed to up-regulate CXCR3 and had an overall reduced capacity to produce IFN-gamma, demonstrating effector T cell differentiation was linked to their migration. To investigate the role of TRIF-dependent cytokines, neutralization studies were performed in wild type mice. Although TNF neutralization reduced T cell numbers, its coneutralization with IL-10 unexpectedly restored the T cells, suggesting the balance between pro- and anti-inflammatory cytokines influences T cell survival rather than their magnitude. To investigate a role for costimulatory molecules, we tested whether the T cell defect in TRIF-deficient mice could be corrected with enforced costimulation. Boosting with a CD40 agonist in addition to LPS restored the effector CD8 T cell response in livers of TRIF-deficient mice while only partially restoring CD4 T cells, suggesting that LPS primes CD8 and CD4 T cell immunity through different mechanisms. Overall, our data support targeting TRIF for vaccines aimed to direct immune responses to nonlymphoid tissues.