Signals mediated by the p38α MAPK have been implicated in many processes required for the development and effector functions of innate and adaptive immune responses. As mice deficient in p38α exhibit embryonic lethality, most analyses of p38α function in lymphocytes have relied on the use of pharmacologic inhibitors and dominant-negative or constitutively active transgenes. In this study, we have generated a panel of low passage p38α+/+, p38α+/-, and p38α-/- embryonic stem (ES) cells through the intercrossing of p38α+/- mice. These ES cells were used to generate chimeric mice by RAG-deficient blastocyst complementation, with the lymphocytes in these mice being derived entirely from the ES cells. Surprisingly, B and T cell development were indistinguishable when comparing chimeric mice generated with p38α+/+, p38α+/-, and p38α-/- ES cell lines. Moreover, proliferation of p38α-/- B and T cells in response to Ag receptor and non-Ag receptor stimuli was intact. Thus, p38α is not an essential component of signaling pathways required for robust B and T lymphocyte developmental, nor is p38α essential for the proliferation of mature B and T cells.