戊型肝炎病毒ORF2片段与ORF3嵌合重组抗原的纯化与免疫学性质  

Purification and Immunological Characterization of Hepatitis E Virus Recombinant Chimeric Antigen Encoded by ORF2 Fragments and ORF3

作　　者：唐浩[1] 马雁冰[1] 杜瑞娟[1] 乐耕耘[1] 李春宏[1] 庄俊英[1] 刘勇[1] 孙茂盛[1] 戴长柏[1] 

机构地区：[1]中国医学科学院 中国协和医科大学 医学生物学研究所分子生物室,昆明650118

出　　处：《中国医学科学院学报》2001年第4期382-385,共4页Acta Academiae Medicinae Sinicae

摘　　要：目的 考察具有多个优势抗原表位区段嵌合表达的戊型肝炎病毒（HEV）重组抗原的免疫学性质。方法 将构建的含有3个不同HEV抗原表位基因(ORF2．1: 6287 ～ 6403nt, ORF2．2: 6743 ～ 7126nt, ORF3)的表达质粒pThioHisORF(2．1＋2．2＋3 )在大肠杆菌中进行异丙基硫代半乳糖苷（IPTG）诱导表达，其表达产物P_(2．1＋2．2＋3 )在8 mol/L尿素下经阳离子柱SP Sepharose FF层析纯化，透析复性。以P_(2．1＋2．2＋3 )免疫家兔和小鼠, 通过ELISA检测实验动物血清及戊型肝炎患者阳性血清IgG抗体，以Western蛋白印迹检测动物免疫血清对杆状病毒系统表达的抗原片段及嵌合抗原P_(2．1＋2．2＋3 )对患者阳性血清的免疫反应性。结果 纯化的抗原纯度达90%以上；免疫动物血清中检测到特异性抗体的滴度分别为1∶25 600（家兔）和1∶12 800（小鼠）以上，抗血清能够特异识别杆状病毒系统表达的HEV抗原片段，P_(2．1＋2．2＋3 )能与患者阳性血清特异反应。结论 嵌合表达的HEV重组抗原P_(2．1＋2．2＋3 )具有良好的免疫原性和免疫反应性，为进一步研究开发HEV诊断试剂盒及基因工程疫苗提供了基础。Objective To study immunological characteristics of recombinant chimeric HEV antigen． Methods Constructed recombinant plasmids pThioHisORF(2．1＋2．2＋3 )，which contains three HEV antigen gene frag- ments(ORF2．1: 6287 ～ 6403nt, ORF2．2: 6743 ～ 7126nt, ORF3)，was transformed into E．coli and induced with IPTG． Expressed product P_(2．1＋2．2＋3 ) existed in inclusion bodies, was purified by denature SP Sepha- rose FF cation exchange chromatography． Rabbits and rats were immunized with renatured P_(2．1＋2．2＋3 )． The level of IgG in sera from experimental animals and clinical patients were examined with P_(2．1＋2．2＋3 )by ELISA． The characteristics of IgG of immunized animals interacted with recombinant antigen expressed by baculovirus system as well as recombinant chimeric antigen interacted with clinical patients sera were evaluated by Western-blotting． Resaults High titer of IgG antibodies, 1∶25 600 in rabbits and 1∶12 800 in rats, were detected after immunized with P_(2．1＋2．2＋3 )． Furthermore, recombinant antigen expressed by baculovirus system was specifically recognized by IgG of experimental animal immunized with P_(2．1＋2．2＋3 ), and the purified recombinant chimeric antigen P_(2．1＋2．2＋3 ) was specifically reacted with the IgG of clinical patients． Conclusions Recombinant chimeric antigen appears a promising strategy for detection of and prevention from HEV infection．

关 键 词：戊型肝炎病毒 重组 纯化 免疫 

分 类 号：R373.21[医药卫生—病原生物学]