抗轮状病毒LLR株G10血清型特异性单克隆抗体的制备与鉴定

目的制备抗轮状病毒(RV)LLR株G10血清型特异性单克隆抗体,并鉴定其特性。方法以纯化的RVLLR株(血清型为G10)为抗原免疫BALB/c小鼠,应用杂交瘤细胞技术筛选分泌G10血清型特异性单抗的杂交瘤细胞株。采用层析法纯化单抗,常规免疫学方法鉴定单抗的特性。结果经筛选获得5G5、1H1和7F6共3株可稳定分泌高效价且具有G10血清型特异性单抗的杂交瘤细胞株。3株细胞分泌的单抗分别为IgG2a/κ、IgG2a/κ和IgG1/κ型;腹水效价均可达1.024×106;分别识别两个抗原位点;相对亲和力依次为1H1﹥7F6﹥5G5;5G5和1H1(细胞上清)均与G1～G4和G6血清型RV抗原无交叉反应;3株杂交瘤细胞分泌的单抗均识别相对分子质量约为34000的VP7蛋白,且均有不同程度的中和活性。结论已成功制备了抗RVLLR株G10血清型特异性单抗,该单抗可用于G10血清型RV的分型检测或LLR毒株的鉴别。

Preparation and Identification of Monoclonal Antibody against Rotavirus LLR Strain Serotype G10

Objective To prepare and identify the monoclonal antibody(McAb)against rotavirus(RV)LLR strain serotype G10.Methods BALB /c mice were immunized with purified RV LLR strain serotype G10,and the hybridoma cell strains secreting specific McAbs were screened.The secreted McAb was purified by chromatography and identified by routine immunological method.Results Three hybridoma cell strains secreting high titer McAbs against RV LLR strain serotype G10 stably,i.e.5G5,1H1 and 7F6,were screened,by which the secreted McAbs were IgG2a /κ,IgG2a /κ and IgG1 /κ respectively.All the secreted McAbs reached titers of 1.024 × 106 in murine ascites,were recognized by VP7 protein with a relative molecular mass of about 34 000,and showed neutralizing activities at different degrees.The McAbs secreted by the three hybridoma cell strains recognized two antigenic epitopes.In the order of relative affinities,the secreted McAbs were 1H1,7F6 and 5G5.Neither 5G5 nor 1H1 showed cross reaction with RV antigen serotypes G1 ～ G4 or G6.Conclusion The McAb against RV LLR strain serotype G10 was successfully prepared,which might be used for G10 serotyping or the identification of LLR strain.