人Lrp蛋白多克隆抗体的制备  

Preparation of human Lrp polyclonal antibody

作　　者：于欣平[1] 宋庆贺[2] 侯立朝[1] 杜可军[3] 熊利泽[1] 陈苏民[2] 陈南春[2] 代忠明[2] 

机构地区：[1]第四军医大学西京医院麻醉科,陕西西安710033 [2]第四军医大学基础部生物化学与分子生物学教研室,陕西西安710033 [3]第四军医大学预防医学系劳动与环境卫生教研室,陕西西安710033

出　　处：《第四军医大学学报》2007年第9期773-775,共3页Journal of the Fourth Military Medical University

基　　金：国家自然科学基金(30170361;30471675)

摘　　要：目的:制备人Lrp蛋白多克隆抗体.方法:克隆全长lrp-cDNA编码序列并进行原核表达与鉴定.用镍离子螯合柱(Ni-NTA)纯化原核表达的全长Lrp蛋白,获得纯度较高的目的蛋白.用纯化后蛋白免疫新西兰大白兔制备多克隆抗体并吸附去除非特异性反应成分.结果:Western印迹结果表明,纯化前后的抗血清在69ku处均检测到目的蛋白条带,说明吸附纯化后的抗体可以与Lrp蛋白特异结合.而纯化后抗体Western印迹结果目的条带只有一条,说明吸附纯化后得到高特异性抗血清,其免疫印迹的效价在10-6以上,有较高免疫印迹滴度.结论:成功制备了高效价高特异性的兔抗人Lrp蛋白的多克隆抗体,为Lrp功能的进一步研究提供了重要的实验工具.AIM： To prepare the human Lrp polyclonal anti- body. METHODS： To research the function of lrp gene further, we cloned total length coding sequence of human lrp-cDNA and made the prokaryotic expression and identification of it. Total length Lrp protein was expressed in E. coli and purified by Ni- NTA chelating column. Polyclonal antibody was prepared by im- munizing New Zealand rabbits with purified protein, then the anti- serum was adsorbed to remove the non-specific reaction compo- nents. RESULTS： Western Blot showed that the band of interest protein at 69 kD was all found before and after the anti-serum was purified, indicating that the adsorbed and purified antibody could combine with Lrp specifically. However, Western Blot also showed that there was noly one band of interest protein after the antibody was purified, indicating that the high specific anti-serum was prepared after adsorption and purification. The immunoblot- ting titer of it was above 10 -6, a very high immunoblotting titer. CONCLUSION： We succeed in preparing the high potency and high specific human Lrp tpolyclonal antibody, offering an imper- tant experimental tool for further reseraching the function of Lrp.

关 键 词：脂多糖类 人脂多糖应答基因 人脂多糖应答蛋白 抗体制备 

分 类 号：Q257[生物学—细胞生物学] Q78