不同基因型戊型肝炎病毒存在多种类型抗原表位

以戊型肝炎病毒(HEV)ORF2重组蛋白p166Us为免疫原制备单克隆抗体(McAbs),采用间接ELISA和免疫印迹法,检测McAbs与不同基因型和亚型HEV重组蛋白p166Bur(Ⅰa型)、p166Pak(Ⅰb型)、p166Mor(Ⅰc型)、p166Mex(Ⅱ型)、p166Us(Ⅲ型)、p166Nz(猪HEV,Ⅲ型)和p166Chn(Ⅳ型)的反应性,采用抗原或抗体竞争ELISA分析p166蛋白与天然HEV颗粒之间抗原表位的关系。结果获得4D3、2E3、11E11、12H5、3A3和1F16株稳定分泌McAbs的杂交瘤细胞株。4D3分泌的McAb与7种p166均发生反应,其与免疫原p166Us的结合可被Ⅰ、Ⅱ、Ⅲ或Ⅳ型天然HEV颗粒或病人血清竞争抑制。2E3、11E11和12H5分泌的McAbs只与p166Us、p166Nz和p166Chn发生反应,它们与p166Us的结合仅能被Ⅲ和IV型病毒或血清所抑制。3A3分泌的McAb只与p166Us及p166Nz结合,1F1分泌的McAb只与p166Us结合,两者均能被Ⅲ型美国株竞争抑制,而Ⅰ、Ⅱ、Ⅳ型不能抑制它们与p166Us的结合。由此可见,不同基因型和亚型HEV ORF2编码蛋白p166上存在多种类型抗原表位,其中包括Ⅰ、Ⅱ、Ⅲ、Ⅳ基因型共同的,Ⅲ、Ⅳ基因型共有的和第Ⅲ基因型特异的等,这些表位与天然HEV颗粒上的抗原表位具有相同的免疫学特征。

Hepatitis E virus of different genotypes contains multiple-type antigenic epitopes

Monoclonal antibodies (McAbs) were prepared against a recombinant protein p166Us derived from US-1 strain of hepatitis E virus (HEV). The immune reactivity of the McAbs to seven p166s derived from different genotypes and subtypes of HEV, which included p166Bur (genotype I a), p166Pak (genotype I b), p166Mor (genotype I c), p166Mex (genotype II), p166Us (genotype III), p166Nz (swine HEV, genotype III) and p166Chn (genotype IV), was tested by an indirect enzyme-linked immunosorbent assay (ELISA) and a Western blotting assay. The immunological relationship between the McAbs and native HEV particles or anti-HEV positive serum samples was analyzed by an antigen-competitive or antibody-competitive ELISA. Totally, six McAb-producible hybridoma cell lines, designated by the name of 4D3, 2E3, 11E11, 12H5, 3A3 and 1F1 respectively, were cloned and obtained in this study. The McAb of 4D3 could react to all of the seven p166 recombinant proteins. This kind of reaction could be inhibited by each of 4 genotypes of native HEV particles or anti-HEV positive serum samples. The McAbs produced by 2E3, 11E11 and 12H5 reacted to p166Us, p166Nz and p166Chn, but did not react to p166Bur, p166Pak, p166Mor and p166Mex. The reaction of McAb of 2E3, as an example of the McAbs of 2E3, 11E11 and 12H5, could be only inhibited by genotype III and IV HEV or anti-HEV positive serum. The McAb of 3A3 could bind to p166Us as well as p166Nz. The McAb produced by 1F1 was reactive to the p166Us only. However, neither of I , II , IV genotype HEV particles or antisera could inhibit both of their reactions to p166Us. The data as mentioned above suggested that there are multiple-type antigenic epitopes such as genotype I, II, III and IV common, III and IV common, and III specific epitopes within HEV ORF2 encoded p166 proteins of different genotypes and subtypes of HEV. Moreover, the antigenic epitopes on recombinant protein p166s and these on native HEV particles possess identical immunological characteristics.