AIM:To construct an eukaryotic superantigen gene expressionvector containing the recombinant gene of SEA and CD80molecule transmembrane region (CD80TM),and to expressstaphylococcus enterotoxin A (SEA) on the membrane ofhepatocellular carcinoma (HCC) cell to form a superantigengene modified tumor vaccine for HCC.METHODS:SEA and linker-CD80TM gene were amplifiedthrough PCR from plasmid containing cDNA of SEA and CD80.Gene fragments were then subcloned into the multiplecloning sites of retroviral vector pLXSN.Recombinant plasmidwas transferred into HepG2 cells mediated with lipofectamine,positive clones were selected in culture medium containingG418.RT-PCR and indirect immunofluorescence studiesconfirmed that SEA was expressed specifically on HCC cellmembrane.INFγ-ELISPOT study demonstrated that SEAprotein was expressed on the membrane of HCC cells.Cytotoxicity of HepG2-SEA primed CTLs (SEA-T) wasanalyzed by ~(51)Cr release assay.T cells cultured with rhIL-2(IL-2-T) were used as control.RESULTS:Restriction digestion and sequence analysesconfirmed the correctness of length,position and orientationof inserted fusion genes.SEA was expressed on the surfaceof HepG2 cells,HepG2-SEA had strong stimulating effect onproduction of HepG2 specific CTL (P<0.001).SEA-T hadenhanced cytotoxicity to HepG2 cells (P<0.05).CONCLUSION:Tumor cell membrane expressed superantigencan be used to reinforce the immune effect of tumor cellvaccine for HCC,which provides a new method of theenhanced active immunotherapy for HCC. AIM: To construct an eukaryotic superantigen gene expression vector containing the recombinant gene of SEA and CD80 molecule transmembrane region (CD80TM), and to expressstaphylococcus enterotoxin A (SEA) on the membrane of hepatocellular carcinoma (HCC) cell to form superantigengene modified tumor vaccine for HCC. METHODS: SEA and linker-CD80 ™ gene were amplified by PCR from plasmid containing cDNA of SEA and CD80. Gene fragments were then subcloned into the multiplecloning sites of retroviral vector pLXSN. Recombinant plasmid was transfected into HepG2 cells mediated with lipofectamine, positive clones were selected in culture medium containing G418.RT-PCR and indirect immunofluorescence studies studies confirmed that SEA was specifically specified on HCC cellmembrane.INFγ-ELISPOT study demonstrated that SEA protein was expressed on the membrane of HCC cells. Cytotoxicity of HepG2-SEA primed CTLs (SEA-T) wasanalyzed by ~ (51) Cr release assay. Cells cultured with rhIL-2 (IL-2-T) were used as control. RESULTS: Restriction digestion and sequence analyzes have confirmed the correctness of length, position and orientation of inserted fusion genes. SEA was expressed on the surface of HepG2 cells, HepG2-SEA had strong stimulating effect onproduction of HepG2 specific CTL (P <0.001) .SEA-T hadenhanced cytotoxicity to HepG2 cells (P <0.05). CONCLUSION: Tumor cell membrane expressed superantigencan be used to reinforce the immune effect of tumor cellvaccine for HCC, which provides a new method of the enhanced active immunotherapy for HCC.