Antigen-Specific Stimulation of CD8+ T-cells by Murine Bone Marrow-Derived Dendritic

The stimulation of antigen-specific T-cell responses is critical for therapeutic cancer vaccines. Extensive efforts have been made to develop nanoparticles as carriers of therapeutic cancer vaccines to antigen presenting cells (APCs) of the immune system. The purpose of this protocol is to use nanoparticles to deliver vaccines to APCs and measure the antigen-specific T-cell responses that are instigated. For the purposes of this assay, the model antigen ovalbumin (OVA) is used in the context of murine cells. The protocol requires the synthesis of nanoparticles that deliver SIINFEKL (OVA257-264), the immunodominant class I peptide derived from OVA, either in its peptide form or as part of a larger molecule (such as the whole OVA protein). CD8+T-cells that recognize SIINFEKL are purified from the transgenic OT-I mouse model, and activated using nanoparticle-treated APCs. The resulting T-cell proliferation, as well as secretion of the cytokines interferon gamma (IFN-γ) and interleukin-2 (IL-2), are used to measure antigen-specific T-cell activation. The results of this protocol may be used to infer the ability of the nanoparticle system to deliver other similar antigens.