In the present study we have investigated the role of the secreted p60 protein from Listeria monocytogenes as an Ag for CD4 T cells. The p60 protein is an abundant extracellular protein that is highly conserved within the members of the genus Listeria. Our results show that L. monocytogenes infection induces a potent p60-specific Th1 immune response. Remarkably, we found that p60-specific Th1 clones mediate significant protection against L. monocytogenes infection. For one p60-specific clone, the peptide epitope was defined. This clone recognized p60 301-312 (EAAKPAPAPSTN) in the context of the H-2Ad molecule. Despite the fact that acquired immunity against L. monocytogenes is primarily mediated by cytotoxic CD8 T lymphocytes, our data clearly demonstrate that secreted bacterial proteins are important CD4 T cell Ags and that Th1 clones specific for a secreted bacterial protein can contribute to the protection against an intracellular pathogen such as L. monocytogenes.