During systemic infection by Listeria monocytogenes the host develops a robust T cell-mediated immune response against the major immunodominant antigens of the pathogen. The enzyme-linked immuno-spot (ELISPOT) test is an accurate and reproducible means of measuring the extent of this T cell response. Here we describe a detailed ELISPOT protocol for measuring an epitope-specific CD8+ T cell-mediated immune response in mice vaccinated with low doses of L. monocytogenes. The basic approach can be easily adapted for the analysis of other vaccination regimes and target epitopes.