基因工程产品中CHO细胞蛋白残余量测定方法的建立  

Establishment of the method for detecting CHO cell protein residue in gene engineering semi finished products

作　　者：邓瑞春[1] 张明伟[1] 白云秀[1] 汪劲松[1] 黄英[1] 毛宁[1] 

机构地区：[1]军事医学科学院瑞得四环生物技术研究所,北京100850

出　　处：《细胞与分子免疫学杂志》1999年第4期292-295,共4页Chinese Journal of Cellular and Molecular Immunology

摘　　要：目的： 建立简便、快速、高灵敏度的基因工程产品中ＣＨＯＤＨＦＲ－ 细胞蛋白的检测方法。方法： 以基因工程用宿主细胞ＣＨＯＤＨＦＲ－ 的全细胞蛋白和该细胞培养上清浓缩蛋白的混合物， 免疫家兔制备抗血清， 并用间接ＥＬＩＳＡ 测定其效价； 用Ｗｅｓｔｅｒｎ ｂｌｏｔ 分析该抗体的特异性； 用该抗体建立直接、间接ＥＬＩＳＡ， 分别测定两种检测方法的动力学曲线、检测范围及灵敏度。结果： 抗ＣＨＯＤＨＦＲ－ 细胞蛋白的抗血清效价为２×１０－ ７ ， 该抗体只与ＣＨＯＤＨＦＲ－ 细胞蛋白起反应， 不与基因工程产品ＥＰＯ 起反应； 用该抗体建立的直接、间接ＥＬＩＳＡ 法检测范围较宽， 灵敏度小于１μｇ／Ｌ。结论： 建立了高灵敏度、高特异性、操作简单、快速， 可检测基因工程产品中ＣＨＯ 细胞蛋白残余量的两种ＥＬＩＳＡ 法。Aim: To establish two simple, fast, sensitive methods for deteting CHODHFR -cell protein residue in the gene engineering semi-finished products Methods: Antiserum was prepared by immunizing the rabbits with mixture of all CHODHFR -cell proteins and concentrated protein in supernatant fluid of CHODHFR -cells; the titers of the antiserum were determined by indirect ELISA, and its specificity was analyzed by Western blot and then the kinetic curve,detection range and sensitivity were determined by direct and indirect methods Results: The titer of the antiserum was as high as 2×10 -7 The antibody reacted only with CHODHFR -cell proteins, but not with the gene engineering product EPO; The linear detection range of the two ELISA methosd was wider, about 4～62 5 μg/L and their sensitivity was lower than 1 μg/L Conclusion: Two ELISA methods established may be used to detect CHO cell protein residue in gene engineering semi finished products

关 键 词：CHO细胞蛋白 ELISA 促红细胞生成素 贫血 

分 类 号：R392－33[医药卫生—免疫学]