兔戊型肝炎病毒ORF2编码蛋白的抗原表位分析  

Analysis of antigenic epitopes of open reading frame 2 encoded protein of rabbit hepatitis E virus

作　　者：王松[1] 戴星[2] 董晨[1] 董敏[1] 梁久红[1] 孟继鸿[1] 

机构地区：[1]东南大学医学院,江苏南京210009 [2]东南大学附属中大医院皮肤科,江苏南京210009

出　　处：《东南大学学报（医学版）》2013年第3期261-266,共6页Journal of Southeast University(Medical Science Edition)

基　　金：国家高技术发展计划(863计划)项目(2006AA02A235);国家自然科学基金资助项目(30671858);江苏省科技支撑计划项目(BE2009664)

摘　　要：目的:研究新发现的兔戊型肝炎病毒(HEV)与所有已知基因型HEV抗原表位的异同,为阐明动物HEV的人畜共患特征及戊型肝炎的诊断提供依据。方法:制备兔HEV ORF2重组蛋白R166(aa452-617),并免疫BALB/c小鼠制备单克隆抗体(McAb);采用间接ELISA法及免疫印迹法,检测McAbs与已知基因型HEV ORF2重组蛋白p166Bur(1型)、p166Mex(2型)、p166US(3型)和p166Chn(4型)的交叉反应性。用获得的杂交瘤细胞制备腹水,利用基于PCR的HEV体外中和试验,检测它们对基因1、4型人HEV和兔HEV的中和活性。结果:获得3株稳定分泌抗-R166的McAbs的杂交瘤细胞株,即1C1、6F9和10D2。其中1C1和6F9分泌的McAbs与已知1、2、3、4基因型p166均结合;10D2分泌的McAb只与R166特异结合,而不与1、2、3、4基因型p166结合。3株McAbs腹水均不能中和基因1、4型人HEV和兔HEV,提示3株McAbs针对非中和性抗原表位。结论:兔HEV与已知1、2、3、4基因型HEV既含有共同的抗原表位,也含有不同的抗原表位,具有不同的抗原特征。Objective： To characterize antigenic epitopes of the newly identified rabbit hepatitis E virus （HEV） and compare the antigenic features between rabbit HEV and all the known genotypes of HEV,which help to clarify zoonotic feature of animal HEV and support evidence of diagnosis and prevention of hepatitis E （HE）. Methods： McAbs were prepared by immunization of mice with recombinant protein R166 （aa452-617） derived from a rabbit HEV strain.The generated McAbs were tested with p166 recombinant proteins derived from HEV genotypes 1,2,3 and 4 using ELISA and Western blotting.They were evaluated for their ability to neutralize genotypes 1,4 and rabbit HEV strains by an in vitro PCR-based HEV neutralization assay. Results： Three hybridoma cell lines screening McAbs 1C1,6F9 and 10D2 against R166 were obtained.Two of the McAbs,1C1 and 6F9,could bind to p166 recombinant proteins derived from HEV genotypes 1,2,3 and 4.However,10D2 reacted to R166 only.All the McAbs did not show neutralizing activity against virus strains of human HEV genotype 1 and 4,as well as rabbit HEV. Conclusion： There are common and different antigenic epitopes between rabbit HEV and all known genotypes of HEV.

关 键 词：兔戊型肝炎病毒 重组ORF2蛋白 抗原表位 单克隆抗体 

分 类 号：R373.21[医药卫生—病原生物学] R392.1[医药卫生—基础医学]