论文部分内容阅读
应用果蝇（ＤＳ２）表达系统，构建了含有乙型肝炎病毒表面抗原（ＨＢｓＡｇ）、摄金蛋白启动子（ＭＴｎ－ｐｒｏｍｏｔｅｒ）的共表达质粒ｐＡＭ－ＨＢｓＡｇ，转染细胞，经克隆，存活细胞株的培养上清液经硫酸铵沉淀、氯化铯（ＣＳＣｌ）密度梯度离心沉淀，获得的抗原用酶联免疫吸附试验（ＥＬＩＳＡ）、放射免疫分析（ＲＩＡ）法检测抗体，免疫吸印法（Ｗｅｓｔｅｒｎｂｌｏｔ）和电泳凝胶银染显色证实分子量为２３０００和２７０００，免疫电镜观察显示表达产物为２２ｕｍ球型颗粒，通过重金属离子（ＣｕＳＯ４、ＺｎＳＯ４）的诱导可增加抗原的表达量。用共表达质粒ｐＡＭ－ＨＢｓＡｇ的ＤＮＡ，注射Ｂａｌｂ／Ｃ小鼠的股四头肌。经ＥＬＩＳＡ、ＲＩＡ检测抗体产生情况，结果免疫后的小鼠经硫酸锌喂养抗体高于普通喂养的小鼠。Ｓｏｕｔｈｅｒｎ杂交证实鼠肌肉细胞存在ＨＢｓＡｇ基因。小鼠免疫接种实验表明，ＤＳ２细胞表达的抗原与直接用ＤＮＡ含有ＨＢｓＡｇ的重组质粒）免疫小鼠均获抗ＨＢｓＡｇ的抗体。
The recombinant plasmid pAM-HBsAg containing hepatitis B virus surface antigen (HBsAg) and superoxide dismutase (MTn-promoter) was constructed by using the DS2 expression system. The transfected cells, cloned and survived cell lines The supernatant was precipitated by ammonium sulfate and centrifuged with CSCl density gradient centrifugation. The obtained antigens were detected by enzyme-linked immunosorbent assay (ELISA), radioimmunoassay (RIA) ) And electrophoresis gel silver staining confirmed that the molecular weight of 23000 and 27000, immunoelectron microscopy showed that the expression product is 22um spherical particles, the expression of antigen can be increased by the induction of heavy metal ions (CuSO4, ZnSO4). The quadriceps of Balb / C mice were injected with DNA that co-expressed plasmid pAM-HBsAg. Antibody production was detected by ELISA and RIA. As a result, the mice immunized with zinc sulfate were higher in the immunized mice than in the normal fed mice. Southern hybridization confirmed the presence of HBsAg gene in murine muscle cells. Mouse immunization experiments showed that the antigen expressed by DS2 cells and recombinant plasmid directly containing DNA containing HBsAg) mice were immunized with anti-HBsAg antibodies.