乙型肝炎病毒重组抗原表位检测敏感性探究.docVIP

乙型肝炎病毒重组抗原表位检测敏感性探究 　 作者：潘云华，周东霞，李嘉琦，陈元鼎 【关键词】 HBV;抗原表位;抗原性;FHV 　　［摘 要］ 目的：在外源抗原表位表达载体系统中构建并表达乙型肝炎病毒(HBV)抗原表位，并对其诊断敏感性进行评价。方法：人工合成编码HBV“a”抗原决定簇表位中24个氨基酸(S124147)的寡核苷酸序列，克隆入外源抗原表位表达载体系统FHVRNA2I3位点，构建重组质粒，在原核pET系统(BL21细胞)中表达。通过ELISA和Western blot检测表达产物的抗原性，并以表达产物为包被抗原,通过ELISA和Western blot检测58份乙肝患者血清抗HBs。结果：嵌和蛋白可与抗HBs特异性结合，其ELISA检测抗HBs阳性率为77.5%，Western blot的为68%，而对照试剂盒的为62%。结论：在外源抗原表位表达系统中表达的HBV重组抗原表位具有良好的抗原性，有诊断应用价值。 　　［关键词］ HBV;抗原表位;抗原性;FHVRNA2载体系统 　　Study on Antigenicity of HBV Recombinant Epitope 　　Abstract：Objective Construction and Expression HBV recombinant epitope in a foreign epitope presenting vector，and evalution of dignostic sensitivity for antiHBs with HBV epitope. Methods Oligonucleotids encoded twentyfour amino acids(aaS124147) which is within “a” antigenic deterninant were synthesized and inserted into a foreign epitopepresenting carrier I3 site (FHVRNA2cDNAI3). HBV epitope was expressed in a prokaryotic cellular system(BL21 cell). The expression product was analyzed and its antigenicity was further studied by ELISA and Wstern blot method with chimeric protein as coating antigen.we used chimeric protein as coating antigen to test antiHBs with ELISA and Wstern blot in 58 seral samples. Results The chimeric protein reacted with antiHBs seral specially.The detection positive ratio of antiHBs is 77.5% and 68%respectively by ELISA and Wstern blot with chimeric protein as coating antigen,the control’s was 62%.Conclusion HBV epitope expressed in a foreign epitopepresenting carrier possessed good antigenicity and diagnostic value. 　　Key words：HBV；Epitope；Antigenicity；FHVRNA2 carrier system 　　乙型肝炎病毒(HBV)是乙型肝炎的病原体，并引起慢性肝炎和肝硬化，而且与肝癌的发生密切相关。迄今，对HBV感染最有力的预防和控制措施，仍是发展疫苗。FHVRNA2载体系统［1］是一个以昆虫小RNA病毒flock house virus外壳蛋白为载体的新型抗原表位表达载体，此系统中表达的载体蛋白可自动组装成病毒样颗粒，使插入的外源抗原表位可以暴露在颗粒的表面，保持天然构象和免疫学特性。本研究拟将HBV a抗原表位插入到FHV外壳蛋白上，并对其抗原性及诊断意义进行了研究。 　　1 材料与方法 　　1.1 细胞 重组质粒DNA大肠杆菌DH5α用于重组质粒pETE DNA的克隆和扩增。大肠杆菌BL21(DE3)用于重组质粒pETE DNA的表达。质粒DNA在含200 μg/ml氨苄青霉素的LB或TB培育中生长，FHVRNA2载体系统即重组pETFHV