膝沟藻毒素GTX1-4单克隆抗体的制备与鉴定

Preparation and identification of monoclonal antibody against Gonyautoxins GTX1-4

研究膝沟藻毒素GTX1-4单克隆抗体的制备方法。利用甲醛法合成GTX1-4人工抗原,通过小鼠免疫和单克隆抗体技术制备GTX1-4单克隆抗体。结果显示,合成了免疫抗原GTX1-4-BSA和检测抗原GTX1-4-KLH,其偶联比分别为10.7∶1与22.4∶1。筛选出3株能稳定分泌GTX1-4单克隆抗体的杂交瘤细胞1-A4、5-E11和5-A11。5-E11、1-A4诱生腹水单克隆抗体的亚型分别为IgG1和IgG2a,效价均达到64000;5-A11诱生腹水单克隆抗体的亚型为IgG1,效价达到128000。5-A11诱生腹水单克隆抗体的灵敏度为6.25μg/mL,特异性较强。本实验成功合成了GTX1-4人工抗原,并获得了能够稳定分泌GTX1-4单克隆抗体的杂交瘤细胞株。

The goal of this study was to develop a method for preparation of monoclonal antibody against Gonyautoxins GTX1-4. GTX1-4 artificial antigen was synthesized by formaldehyde method, and GTX1-4 monoclonal antibody was prepared by mouse immunity and monoclonal antibody technology. The results showed that the conjugated ratios of immunizing antigen GTX1-4-BSA and detected antigen GTX1-4-KLH were 10.7∶1 and 22.4∶1, respectively. Three hybridomas cell lines, 1-A4, 5-A11 and 5-E11, which could stably secret GTX1-4 monoclonal antibody were screened. The subtypes of monoclonal antibodies in ascites induced by 5-E11 and 1-A4 were IgG1 and IgG2 a, and the titers of them were up to 64000. The subtype of monoclonal antibody in ascites induced by 5-A11 was IgG1, and its titer could be 128000. The sensitivity of monoclonal antibody in ascites induced by 5-A11 was 6.25 μg/mL, and this antibody has a strong specificity. In the study, consequently, GTX1-4 artificial antigen was successfully synthesized, and the hybridomas cell lines which could stably secret GTX1-4 monoclonal antibody was obtained.