Abstract. T cell-mediated antitumor immunity is a cellular immune response that requires two signals. The dendritic cell (DC) has been considered as the most efficient antigen-presenting cell (APC). It plays essential roles in the induction, regulation and maintenance of antitumor immunity in humans. The 4-1BB/4-1BB ligand (4-1BBL) pathway plays crucial roles in immune response, tumor immunity and autoimmune diseases through transduction of T cell co-stimulatory signals. The aim of this study was to generate the preparation protocol for a DC vaccine transfected with gastric cancer cell total ribonucleic acid (RNA) carrying the 4-1 BBL gene in vitro and to investigate its antitumor effects in murine forestomach carcinoma (MFC). The vaccine was prepared by transfecting MFC total RNAs carrying the 4-1BBL gene into the DCs that were isolated from 615 mouse bones. The T cell proliferation rate in the MFC/4-1BBL/DC group was higher than that in the DC group. The tumor cell kill rate in the MFC/4-1BBL/DC group was higher than that in the DC group. ELISA analysis showed that IL-12 and IFN-γ in the MFC/4-1BBL/DC group were more highly expressed compared to the other group. Collectively, our data demonstrate that the DC vaccine transfected with gastric cancer cell total RNA carrying the 4-1BBL gene has a stronger ability to kill gastric cancer cells through promoting T cell proliferation and enhancing the ability of cytotoxic T lymphocytes (CTLs) to kill gastric carcinoma cells and to secrete IL-12 and IFN-γ. Our results provide an effective therapeutic strategy for treating gastric cancer using a DC vaccine.