水泡性口炎病毒单克隆抗体的制备与鉴定

将水泡性口炎病毒（VSV）经差速离心和蔗糖密度梯度离心法进行纯化后,以纯化的VSV作为免疫原免疫8～10周龄雌性BALB/c小鼠,取其脾细胞与SP2/0骨髓瘤细胞进行融合,经间接ELISA方法筛选能稳定分泌抗VSV单克隆抗体的杂交瘤细胞株,并对制备出的抗VSV单抗的特异性、抗体亚类等生物学特性进行鉴定。结果显示,试验成功筛选出2株能稳定分泌抗VSV单克隆抗体的杂交瘤细胞株,分别命名为1A2、4C3。ELISA鉴定结果表明,2株单抗均能特异性地与VSV结合,而与口蹄疫病毒（FMDV）、猪水泡病病毒（SVDV）均不发生交叉反应;诱生小鼠腹水产生的抗体效价可达1∶25 600～1∶51 200。杂交瘤细胞染色体核型鉴定结果显示,杂交瘤细胞染色体数为95～105,均高于2个亲本细胞的染色体数目,说明这2株细胞是两者的杂交产物。抗体亚类鉴定结果显示,所获得2株单抗1A2、4C3均为IgG1。Western blot分析结果表明,1A2可识别VSV G蛋白。2株抗VSV单克隆抗体的成功制备将为VSV快速检测方法的建立以及检测试剂的研制等奠定基础。

Preparation and identification of monoclonal antibody against vesicular stomatitis virus

Vesicular stomatitis virus(VSV) was purified by differential and sucrose density gradient centrifugations,then 8-10 weeks old female BALB/c mice were immunized with the purified VSV.McAbs against VSV were produced by fusing SP2/0 myeloma cells with spleen cells of BALB/c mice immunized with purified VSV and were screened by indirect ELISA.Then specificity,sub-class of antibodies and other biological characteristics of McAb against VSV was identified.The results showed two hybridoma cell strains secreting McAbs against VSV were screened,named 1A2 and 4C3.ELISA identification results showed that the two McAbs could specifically react to VSV but not cross react to foot and mouth disease virus(FMDV),swine vesicular disease virus(SVDV).The ELISA titers of 2 McAbs in mouse ascites were 1∶25 600-1∶51 200.Hybridoma karyotype indentification results showed that the number of hybridoma chromosome was 95-105,higher than the number of two parental cell chromosome,indicating that two cells were the product of hybridization between the two parental cells.Identification of antibody subclasses showed that the two monoclonal antibodies 1A2,4C3 were IgG1.Western blot analysis showed that 1A2 recognized VSV G protein.The McAbs against VSV was successfully prepared,which laid a foundation of developing a rapid determination method for VSV.