欢迎访问过程工程学报, 今天是

›› 2008, Vol. 8 ›› Issue (4): 789-793.

• 生化工程专栏 • 上一篇    下一篇

cTnI特殊位点抗原的重复表达、免疫及其抗体效价检测

时丽丽 李强 袁其朋   

  1. 北京化工大学生命科学与技术学院 清华大学化学工程系 北京化工大学生命科学与技术学院
  • 收稿日期:2008-02-13 修回日期:2008-04-30 出版日期:2008-08-20 发布日期:2008-08-20
  • 通讯作者: 时丽丽

Expression of cTnI Specific Antigen Sites and Antibodies Detection

SHI Li-li LI Qiang YUAN Qi-peng   

  1. College of Life Science and Technology, Beijing University of Chemical Technology Department of Chemical Engineering , Tsinghua University School of Life Science and Technology, Beijing University of Chemical Technology
  • Received:2008-02-13 Revised:2008-04-30 Online:2008-08-20 Published:2008-08-20
  • Contact: SHI Li-li

摘要: 构建了含人cTnI蛋白稳定区段2个特殊抗原决定簇位点的pMAL-Antigen质粒. 利用BamHI/BgLII两限制性内切酶的同尾酶作用进行连接,在pMAL-p2x质粒的MBP(麦芽糖结合蛋白)之后接入了上述5个相同的片段. 此质粒在大肠杆菌E. coli BL21 (DE3)中经1.0 mmol/L IPTG诱导表达出的融合蛋白MBP-Antigen经Amlyose-resin亲和层析柱纯化后免疫BABL/C小鼠,成功获得了能抗人cTnI和抗牛cTnI的抗血清.

关键词: 人cTnI, 特殊位点, 麦芽糖结合蛋白, Amylose-resin亲和层析, 抗体

Abstract: Two specific epitope sites of human cardiac Troponin I (hcTnI), 38aa~52aa and 87aa~97aa, which can stably exist in the blood serum of acute myocardial infarction (AMI) patients, were selected as objective gene sequences and isolated by 5 lysines with low immunogenicity. Two specific epitopes and lysines constituted AA gene sequence. Plasmid pMAL-Antigen consisting of 5AAs after maltose binding protein (MBP) was constructed by two restriction endonucleases, BamHI and BgLII. The pMAL-Antigen plasmid was diverted into E. coli BL21(DE3), and MBP-Antigen fusion protein was induced by 1.0 mmol/L IPTG in E. coli BL21(DE3). The MBP-Antigen fusion protein was purified by Amylose-resin. The result showed that protein antigen of MBP-Antigen fusion protein was partially reduced from C end. The purity of MBP-Antigen fusion protein was over 90%. The hyper-purified MBP-Antigen fusion protein was injected into BALB/C mice and antibodies were detected by ELISA. These antibodies could be applied in the diagnosis of AMI and myocardial damage.

Key words: human cTnI, specific epitope sites, maltose binding protein, Amylose-resin, antibody

中图分类号: