摘要
目的 以免疫亲和层析方法纯化重组人促红细胞生成素 (rhEPO)。方法 以rhEPO作为抗原 ,免疫Balb/C小鼠 ,制备抗rhEPO单克隆抗体 (mAb)。以纯化的抗rhEPOmAb 2F12与预活化的Sepharose 4B偶联 ,制成抗体亲和层析柱 ,纯化rhEPO。结果 经筛选共获得 4株可分泌抗rhEPOmAb的杂交瘤细胞株 ,分别为 :1A8、2F12、3D4和 3F10。亲和层析纯化的rhEPO经SDS PAGE显示单一条带 ;2g凝胶制备的层析柱一次层析可获得 1.5mg高纯度的rhEPO。利用纯化的mAb建立的ELISA法 ,用于检测rhEPO的蛋白含量可达ng水平。结论 免疫亲和层析法纯化rhEPO的效率高、纯度好 ,用纯化的mAb建立的ELISA法具有灵敏度高。
Aim To purify recombinant human erythropoietin(rhEPO) by immunoaffinity chromatograghy. Methods rhEPO was used as antigen to immunize Balb/c mice for preparing mAbs against rhEPO. Purified mAb 2F12 was coupled with activited sepharose 4B to prepare the immunoaffinity chromatography columu for purification of rhEPO. Results Four hybridoma cell lines secreting mAbs against rhEPO were obtained, namely, 1A8、 2F12、 3D4 and 3F10. A single band be exhibited on SDS PAGE gel with a yield of 90%. By using sandwich ELISA established with these mAbs for detection of rhEPO, Sensitivity of the ELISA reached ng level. Conclution rhEPO purified by immunoaffinity chromatograghy have high efficien good purify and the ELISA are sensitiveand and stabile.
出处
《细胞与分子免疫学杂志》
CAS
CSCD
北大核心
2002年第4期408-411,共4页
Chinese Journal of Cellular and Molecular Immunology
