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目的制备抗人IL-17单克隆抗体,并鉴定其中和活性。方法用hIL-17作为免疫和检测抗原,用间接ELISA法筛选分泌抗人IL-17抗体的杂交瘤细胞株,并对抗体进行中和活性鉴定。结果获得1株稳定分泌抗人IL-17单克隆抗体的杂交瘤细胞株,所分泌的单抗类型重链为IgG2b,轻链为κ;该株杂交瘤细胞腹水效价为1∶8.192×105;传30代及液氮中保存6个月,抗体效价稳定;Western Blot检测证明该单抗与人IL-17蛋白特异地结合;单抗亲和常数为8.192×10-9 mol/L;ELISA及Real-time-PCR检测证明该单抗能够有效地阻断人IL-17刺激Hela细胞产生IL-6的作用。结论所制备的抗人IL-17单克隆抗体具有高度的特异性、稳定性及中和活性,为针对IL-17为靶点的抗体药物的开发奠定了基础。
Objective To prepare anti-human IL-17 monoclonal antibody and identify its neutralization activity. Methods The anti-human IL-17 antibody-secreting hybridoma cell line was screened by indirect ELISA with hIL-17 as an immunogen and antigen. The neutralizing activity of the antibody was evaluated. Results A hybridoma cell line stably secreting anti-human IL-17 monoclonal antibody was obtained. The secreted mAb heavy chain was IgG2b and the light chain was κ. Ascites titer of the hybridoma was 1: 8.192 × 105 ; After 30 passages and liquid nitrogen storage for 6 months, the antibody titer was stable; Western Blot showed that the McAb specifically bound with human IL-17 protein; the affinity constant of monoclonal antibody was 8.192 × 10-9 mol / L; ELISA and Real-time-PCR showed that the McAb could effectively block the effect of human IL-17 on IL-6 production in Hela cells. Conclusion The prepared anti-human IL-17 monoclonal antibody has high specificity, stability and neutralization activity, which lays the foundation for the development of anti-IL-17 antibody.