To evaluate the usefulness of monocyte-derived dendritic cells transfected with tumor antigen mRNA for dendritic cell-based antitumor immunotherapy, we attempted to generate antigen-specific cytotoxic T cells by priming lymphocytes with monocyte-derived dendritic cells transfected with in vitro-transcribed tumor antigen mRNA. Mature monocyte-derived dendritic cells were generated from microbeads-separated CD14(+) cells by culturing with GM-CSF/IL-4 for 7 days and with TNF-alpha, IL-1alpha, IL-6, and PGE(2) for the last one day. Monocyte-derived dendritic cells, lymphocytes, and target cells, which were positive for HLA-A24, were used in the present study. Although lymphocytes prestimulated with untransfected monocyte-derived dendritic cells did not possess the cytotoxic ability against the target cells in a (51)Cr-release cytotoxicity assay, lymphocytes primed with tumor antigen RNA-transfected monocyte-derived dendritic cells were cytotoxic against the tumor antigen-expressing cells but not against the target cells without the expression of the antigen. The cytotoxic ability of the lymphocytes was blocked by the addition of antibodies against MHC class I but not by antibodies against MHC class II. These findings revealed that monocyte-derived dendritic cells transfected with WT1 or SART1 mRNA are able to induce tumor antigen-specific cytotoxic T cells and applicable for antitumor dendritic cell-based cellular immunotherapy.