抗银杏二萜内酯葡胺注射液蛋白单克隆抗体制备.PDFVIP

·2828 · 中草药 Chinese Traditional and Herbal Drugs 第 44 卷 第 20 期 2013 年 10 月 抗银杏二萜内酯葡胺注射液蛋白的单克隆抗体制备 1, 2, 3 4 1, 2, 3 4 1, 2, 3* 李 芳 ，赵 妮 ，王振中 ，秦爱建 ，萧 伟 1. 江苏康缘药业股份有限公司，江苏 连云港 222001 2. 中药制药过程新技术国家重点实验室，江苏 连云港 222001 3. 江苏省企业院士工作站，江苏 连云港 222001 4. 扬州大学兽医学院，江苏 扬州 225009 摘 要：目的 制备抗银杏二萜内酯葡胺注射液蛋白的单克隆抗体。方法 将银杏叶总蛋白进行二维电泳，取蛋白点进行质 谱分析并合成多肽。利用多肽作为免疫原免疫 BALB/c 小鼠，取其脾细胞与 SP2/0 细胞进行融合，通过间接 ELISA 方法筛 选抗银杏二萜内酯葡胺注射液蛋白的单克隆抗体。结果 5 株单克隆抗体亚类鉴定均为IgM，轻链为Kappa 链。Western blotting 实验证实单抗均能与银杏叶蛋白反应。结论 单克隆抗体的制备为快速检测银杏二萜内酯葡胺注射液蛋白的方法建立奠定 了基础。 关键词：银杏二萜内酯葡胺注射液；银杏叶；蛋白；单克隆抗体；免疫原 中图分类号：R285.5 文献标志码：A 文章编号：0253 - 2670(2013)20 - 2828 - 05 DOI: 10.7501/j.issn.0253-2670.2013.20.008 Preparation of monoclonal antibody against proteins of ginkgo diterpene lactones meglumine injection 1, 2, 3 4 1, 2, 3 4 1, 2, 3 LI Fang , ZHAO Ni , WANG Zhen-zhong , QIN Ai-jian , XIAO Wei 1. Jiangsu Kanion Pharmaceutical Co., Ltd., Lianyungang 222001, China 2. State Key Laboratory of Pharmaceutical Process New-Tech for Chinese Medicine, Lianyungang 222001, China 3. Enterprises Academician Workstations in Jiangsu Province, Lianyungang 222001, China 4. College of Veterinary Medicine, Yangzhou University, Yangzhou 225009, China Abstract: Objective To obtain the monoclonal antibodies against the protein of ginkgo diterpene lactones meglumine injection. Methods After total proteins of ginkgo leaf running two-dimensional electrophoresis, the protein spots were digged and analyzed by mass spectrometry (MS). According to the result of MS, the polypeptide was synthesized and used to immunize the BALB/c mice. The indirect ELISA was utilized to develop monoclonal antibody after cell fusion between SP2/0 cells and spleen cells from the immunized BALB/c mic