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为获得针对鲤春病毒血症病毒(SVCV)特异性的单克隆抗体(MAb),以纯化的SVCV为抗原,免疫BALB/c小鼠。将免疫鼠的脾细胞与SP2/0骨髓瘤细胞融合,采用间接ELISA法筛选获得4个能稳定分泌抗SVCV MAb的杂交瘤细胞株;4个杂交瘤细胞制备腹水的MAb效价为1∶160000～1∶640000。亚型鉴定结果表明,这些MAb分属2个亚型(1F1、3E1,IgG2a;3F5、4F9,IgG1),轻链均为Κ链。Western blot分析显示,MAb1F1、3F5、4F9能特异性地识别SVCV的N蛋白(47ku),3E1能特异性地识别SVCV的G蛋白(69ku)。采用相加ELISA法对抗原表位分析结果显示,1F1、3F5、4F9可能识别相同的表位,3E1则识别不同的表位。间接免疫荧光试验结果显示4株MAb均能对染毒病灶产生特异性的荧光染色。这些MAb的制备为SVCV免疫学检测方法的建立奠定了基础。
BALB / c mice were immunized with purified SVCV as antigen for obtaining monoclonal antibody (MAb) specific for the carp virus viremia virus (SVCV). The spleen cells of immunized mice were fused with SP2 / 0 myeloma cells, and four hybridoma cell lines that can stably secrete anti-SVCV MAb were screened by indirect ELISA. The MAb titers of 4 hybridoma cells producing ascites were 1:160000 ~ 1: 640000. Subtype identification results showed that these MAb belong to two subtypes (1F1, 3E1, IgG2a; 3F5, 4F9, IgG1), light chains are Κ chain. Western blot analysis showed that MAb1F1, 3F5 and 4F9 could specifically recognize the N protein of SVCV (47ku), and 3E1 could specifically recognize the G protein of SVCV (69ku). Addition of ELISA method for antigen epitope analysis showed that 1F1, 3F5, 4F9 may recognize the same epitope, 3E1 then identify different epitopes. Indirect immunofluorescence test results showed that four strains of MAb can produce specific fluorescent staining of the infected lesions. The preparation of these MAb lays the foundation for the establishment of SVCV immunological detection method.