L-selectin is a homing receptor that mediates the selective attachment of leukocytes to specialized high endothelial venules. To study the potential role of L-selectin in immune responses in intact mice, we generated L-selectin-deficient mice by gene targeting. L-selectin-deficient mice are defective in cutaneous delayed-type hypersensitivity (DTH) responses when tested after conventional intervals of immunization (4 d). Primary T cell proliferative responses and cytokine production (interleukin [IL] 2, IL-4, and interferon gamma) were also compromised when tested after 5 d of immunization, indicating that L-selectin is important for the immune response to antigens. In contrast, after more prolonged immunization protocols (9 d), normal responses were observed, suggesting that L-selectin-independent compensatory mechanisms exist. Interestingly, humoral responses of L-selectin-deficient mice to keyhole limpet hemocyanin are indistinguishable from wild-type control mice, implying that L-selectin plays no rate-limiting role in T cell help of B cell function. Thus, our results suggest that L-selectin plays an important role in the generation of primary T cell responses but may not be essential for humoral and memory T cell responses. L-selectin does not appear to be rate limiting for the events leading to antigen-driven neutrophil recruitment, since normal DTH responses are obtained at late time points after immunization.