抗人精子膜80KD蛋白单克隆抗体的制备及其特性鉴定

Preparation and Characteristics of McAb Against 80KD Protein of Human Sperm Membrane

目的 :探索避孕疫苗的制作并为不育病人提供抗精子抗体的诊断方法。方法 :用 0 5 %NP -4 0提取经SDS-PAGE电泳分离纯化获得的人精子膜 80KD蛋白免疫BALB/C小鼠 ,取其脾细胞与SP2 /O小鼠骨髓瘤细胞融合 ,获得 1 2株阳性单克隆细胞株 ,并对其中 4株进行了初步鉴定。结果 :免疫双散法结果显示 ,3株为IgG型 ,1株为IgM型 ;免疫荧光定位试验显示 4株单克隆抗体主要与人精子顶体后、颈和尾中段结合 ;Westernblot试验结果显示 ,4株单克隆抗体均识别人精子80KD蛋白。结论 :4株单克隆抗体对人精子膜 80KD蛋白是特异的。

Objective:To explore the preparation of contraceptive vaccine and to diagnose the sterile patients with the antisperm McAb.Methods:The 80KD protein of human sperm membrane was extracted with 0.5% NP-40 and subsequently purified with SDS-PAGE.Spleen cells of mouse immunized with 80KD protein were fused with Sp2/0 myeloma cells and twelve hybridoma clones obtained.Out of 12 strains,4 McAbs were preliminarily identified by agar double diffusion test,immunoflurescence test and western blot test respectively.Results:All four McAbs recongnized 80KD protein and bound specifically to post-acrosome,neck and tail midpiece of sperm.Three McAbs were IgG type,one IgM type.Conclusion:The 4 McAbs obtained show specific to 80KD protein of human sperm membrane.