粒细胞集落刺激因子基因克隆、表达及免疫检测

G-CSF GENE CLONING,EXPRESSION AND IMMUNOTESTIFICATION

目的：获得纯净的抗Ｇ－ＣＳＦ蛋白的抗体，用以鉴定、检测和纯化人Ｇ－ＣＳＦ。方法：将经过改造的粒细胞集落刺激因子（Ｇ－ＣＳＦ）基因重组到ＰＷＲ－４５０－１载体上得到重组质粒，转化大肠杆菌，抽提纯化ＰＷＲ－４５０－ＧＣＳＦ融合蛋白，作为抗原用以免疫兔子，获得抗血清，并对抗血清进行初步纯化，检测其效价和特异性程度。结果：用ＥＬＩＳＡ方法和ＷｅｓｔｅｒｎＢｌｏｔ方法检测抗血清发现本实验所获得的抗血清和融合蛋白及ＰＷＲ－４５０－１均有专一性反应，有一定的特异性。

In order to acquire enough pure antibody against G-CSF for future analysis,we reconstructed prokaryotic expression vector PWR-450-1 which can express G-CSF fusion protein.The expression condiotion was optimized ,and the acquired protein was used to immune rabbit.The Antiserum was further purified and checked for its specificity.Elisa and Westernblot experiment showed that antibody against G-CSF was specific.