鱼肠道弧菌单克隆抗体的制备及应用

Preparation and characterization of monoclonal antibodies against Vibrio ichthyoenteri

以鱼肠道弧菌Vibrio ichthyoenteri为抗原免疫小鼠,用PEG法将小鼠脾细胞与SP2/0骨髓瘤细胞融合,分别采用酶联免疫吸附(ELISA)技术和间接免疫荧光抗体(EFAT)技术筛选强阳性杂交瘤细胞株,亚克隆阳性细胞株,获得5株抗鱼肠道弧菌单克隆抗体(1C6、1D8、2D7、2D10、2E3)。Western blotting分析结果显示,单抗2D10与相对分子质量为44 200、36 700的鱼肠道弧菌蛋白发生特异性结合;用流式细胞术(FCM)检测单抗强度,结果显示阳性荧光比例为13.63%,阴性对照荧光比例不足1%;人工感染牙鲆试验结果表明,运用该单抗能够建立鱼肠道弧菌的快速诊断方法。

Five monoelonaI antibodies （MAbs） （ 1 C6,1D8,2D7,2D10,2E3 ） against Vibrio ichthyoenteri as a pathogen in marine animals were produced from hybridoma cells through Enzyme Linked Immunosorbent Assay （ELISA） and Indirect Immunofluorescenee Assay Test （IFAT） by immunised Balb/C mice. The western blotting revealed that the protein of V. ichthyoenteri was shown to have molecular weight of 44 200 and 36 700, and the flow- ing cytometry （FCM） analysis showed that the MAb 2D10 reacted with positive cells had positive percent of 13.63%, and negative percent of less than 1%. Artificial infection experiment proved that the application of mono- clonal antibody provided a method for the early diagnosis of V. ichthvoenteri.