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将抗癌胚抗原单链抗体基因与锰 超氧化物歧化酶基因融合 (ScFv Mn SOD )插入昆虫杆状病毒供体质粒pFastBacHTb中 ,经大肠杆菌DH1 0Bac体内转座 ,产生重组杆状病毒pBacHTb Mn SOD ScFv。将其转染粉纹夜蛾Tn 5B1 4细胞 ,经扩增后在细胞内进行表达。SDS PAGE分析结果表明 ,融合基因得到高效表达 ,其表达产物相对分子质量为 40 0 0 0单体和 1 60 0 0 0左右的四聚体。Western印迹分析结果 ,以 6×His单克隆抗体为一抗进行蛋白印迹在相对分子质量 40 0 0 0单体和 1 60 0 0 0四聚体处可见表达条带 ,放射免疫分析表明重组杆状病毒表达ScFv Mn SOD融合蛋白能与CEA有较高的结合力 ,并且此融合蛋白具有特异SOD酶活性 ,酶比活可达 32 6 5u/mg。
The anti-CEA single-chain antibody gene and manganese superoxide dismutase gene fusion (ScFv Mn SOD) into the insect baculovirus donor plasmid pFastBacHTb, E. coli DH1 0Bac transposition in vivo to produce recombinant baculovirus pBacHTb Mn SOD ScFv. The recombinant plasmids were transfected into the Trichinella spiralis Tn 5B1 4 cells and then expressed in the cells after amplification. SDS PAGE analysis showed that the fusion gene was highly expressed, and the relative molecular mass of the expressed product was about 40,000 and tetramers about 160,000. Western Blot analysis showed that 6 × His monoclonal antibody was used as a Western blot in the expression of the relative molecular mass of 40 0 0 0 monomer and 1 60 0 0 0 tetramer expression bands can be seen by radioimmunoassay recombinant rod The virus expressed ScFv Mn SOD fusion protein with CEA higher binding capacity, and the fusion protein has a specific SOD activity, enzyme specific activity up to 32 6 5u / mg.