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Summary: We evaluated γ-irradiated Listeria monocytogenes as a killed bacterial vaccine, testing the hypothesis that irradiation preserves antigenic and adjuvant structures destroyed by traditional heat or chemical inactivation. Irradiated Listeria monocytogenes (LM), unlike heat-killed LM, efficiently activated dendritic cells via Toll-like receptors and induced protective T cell responses in mice. Like live LM, irradiated LM induced Toll-like-receptor-independent T cell priming. Cross-presentation of irradiated listerial antigens to CD8+ T cells involved TAP- and proteasome-dependent cytosolic antigen processing. These results establish that killed LM can induce protective T cell responses, previously thought to require live infection. γ-irradiation may be potentially applied to numerous bacterial vaccine candidates, and irradiated bacteria could serve as a vaccine platform for recombinant antigens derived from other pathogens, allergens, or tumors. [Copyright &y& Elsevier]