微量免疫法制备小鼠抗人补体C9抗血清

Preparation of antiserum to human C9 with a new immunization method using trace amount of immunogen

目的 建立一种简便有效的免疫方法 ,制备抗人补体C9抗血清。方法 将鉴定好纯度的免疫原稀释至恰当浓度 ,以硝酸纤维素膜吸附抗原 ,将之埋入小鼠背部皮下 ,并再次免疫 1～ 2次。间接ELISA法鉴定抗血清效价 ,免疫印迹实验鉴定抗血清的免疫反应特异性。结果 间接酶联免疫吸附实验滴定抗血清的效价约为 1× 10 -5,抗血清对正常人血清成分进行的免疫杂交实验得到特异性的C9蛋白条带。结论 以硝酸纤维素膜吸附抗原的微量免疫方案获得特异性和效价均佳的免疫效果。

Objective To establish a simple and effective immunization protocol to prepare antiserum to human C9 in mice. Methods After characterized purity of C9 with high performance capillary electrophoresis, the immunogen was diluted into appropriate concentration and incubated with NC membrane. Mice were immunized with NC membrane loading C9 protein. The titer of antiserum was detected with undirected enzyme linked immunosorbent assay and the specificity was shown with western blotting. Results The titer of antiserum is about 1×10 -5 in ELISA. Immunological blotting using the antiserum as first antibody showed specific C9 band in sample of normal human serum. Conclusion The immunization method using NC membrane as an immunogen carrier has succeed in getting antiserum with high titer and specificity.