抗磷酸酪氨酸抗体的制备

目的；制备抗磷酸酪氨酸抗体。方法：以EDC（碳化二亚胺）交联法合成磷酸酪氨酸牛血清白蛋白（P－Tyr－BSA）；以P－Tyr－BSA免疫新西兰免，抗血清经硫酸该盐析和DEAE纤维素柱层析纯化；以琼脂糖双扩法测定抗体效价。结果：得到抗血清效价为1：32，抗血清与牛血清蛋白（BSA）交叉反应的效价为1：2。纯化的抗体与磷酸酪氨酸牛血清白蛋白、磷酸酪氨酸卵清蛋白、磷酸丝氨酸牛血清白蛋白、磷酸苏氨酸牛血清白蛋白反应效价分别为1：32、1：16、1：2、1：2，所得抗体蛋白量为120mg。纯化的抗体在醋酸纤维素薄膜上电泳为单一区带。结论：用此法制备得的抗体专一性高，交叉反应低，抗体量大。

PREPARATION OF ANTIPHOSPHOTYROSINE ANTIBODY

Objective:To get antiphosphotyrosine antibody. Methods: Phosphotyrosine bovine serum albumin complex (P-Tyr-BSA) was synthesized by EDC 1-(3-Dimethylaminopro pyl ) - 3-ethylcarbodiimidehydrochloride] cross binding. P- Tyr -BSA complex was used as an antigen to immunize rabbit for fourtimes. The antibody titration was performed by using the agar-precipitation double-diffusion method.Results:The antiserum titration was 1: 32. After ammonium sulfate precipitation and DEAE colunmchromatograph,the antibody was purified. The antibody show only one band in the electrophoresis oncellulose acetate membrane. The titrations of purified antibody were 1: 32 with P-Tyr-BSA, 1: 16 withphosphotyrosine oocyte albumin (P-Tyr-OA ). The purified antibody had less cross response to P-Thr-BSA or P-Ser-BSA(tetration 1: 2). The total proteins of the antibody was 120 mg. Conclusion:Thismethod is simple and can get large quantity of the specific antibody