Murine in vivo CD8⁺ T Cell Killing Assay

Antigen-specific killing ability of effector CD8⁺ T cells is critical for protective immunity against infection. Here, we describe in vivo cytotoxic T cell assay to examine effector function of antigen-specific CD8⁺ T cells. Mice infected with Listeria monocytogenes (L. monocytogenes) expressing chicken ovalbumin as a model antigen mount ovalbumin-specific CD8⁺ T cell responses. Effector CD8⁺ T cell function in vivo is determined by mixed transfer of OVA peptide-pulsed target cells with control target cells into the previously immunized mice. Difference in CFSE expression levels clearly marks two distinct populations: Antigen-pulsed target cells-CFSE^low vs. unpulsed target cells-CFSE^hi. The frequencies between antigen-pulsed target cells and control target cells are used as readouts of antigen-specific killing.