高等学校化学学报 ›› 2020, Vol. 41 ›› Issue (7): 1552-1558.doi: 10.7503/cjcu20200078

• 有机化学 • 上一篇    下一篇

磷酰化组氨酸特异性抗体的制备与应用

李容庆1,2,刘佳璐1,2,范若辰3,白鹏阳1,2,张丽影1,*(),权春善1,2,*()   

  1. 1.大连民族大学生命科学学院, 大连 116600
    2.生物技术与资源利用教育部重点实验室, 大连 116600
    3.大连理工大学生命科学学院, 大连 116024
  • 收稿日期:2020-02-17 出版日期:2020-07-10 发布日期:2020-03-23
  • 通讯作者: 张丽影,权春善 E-mail:zhangly@dlnu.edu.cn;mikyeken@dlnu.edu.cn
  • 基金资助:
    国家自然科学基金(21272031);国家自然科学基金(21372037);辽宁省自然科学基金(20180550977);中央高校自主科研基金(wd01190)

Preparation and Application of Specificity Phosphohistidine Antibody

LI Rongqing1,2,LIU Jialu1,2,FAN Ruochen3,BAI Pengyang1,2,ZHANG Liying1,*(),QUAN Chunshan1,2,*()   

  1. 1. College of Life Science, Dalian 116600, China
    2. Key Laboratory of Biotechnology and Bioresources Utilization, Ministry of Education, Dalian Minzu University, Dalian 116600, China
    3. College of Life Science, Dalian University of Technology, Dalian 116024, China
  • Received:2020-02-17 Online:2020-07-10 Published:2020-03-23
  • Contact: Liying ZHANG,Chunshan QUAN E-mail:zhangly@dlnu.edu.cn;mikyeken@dlnu.edu.cn
  • Supported by:
    † National Natural Science Foundation of China(21272031);National Natural Science Foundation of China(21372037);Natural Science Foundation of Liaoning Province, China(20180550977);Fundamental Research Funds for the Central Universities of China(wd01190)

摘要:

以4-碘吡唑和亚磷酸二乙酯为初始原料, 经3步反应合成了新型的磷酰化组氨酸结构类似物, 并以其为半抗原制备了磷酰化组氨酸特异性抗体(Anti-pHis). 采用酶联免疫吸附(ELISA)、 斑点印迹(Dot blot)和免疫印迹(Western blot)实验等方法对Anti-pHis的特异性进行了检测. 结果表明, Anti-pHis对组氨酸磷酰化蛋白表现出优异的亲和力和选择性识别能力, 并可应用于微生物细胞裂解液中组氨酸磷酰化蛋白的检测.

关键词: 半抗原, 磷酰化组氨酸抗体, 特异性, 亲和力

Abstract:

A novel phosphorylated histidine analogue was synthesized using 4-iodopyrazole and diethyl phosphite as raw materials and characterized by nuclear magnetic resonance spectroscopy(NMR). It was used as hapten to prepare phosphohistidine antibody(Anti-pHis). Enzyme-linked immunosorbent assay, dot blot and western blot methods were used to detect the specificity and sensitivity of the Anti-pHis. The results showed that the Anti-pHis had excellent avidity and sensitivity for histidine phosphorylated protein, and the Anti-pHis was successfully applied to the detection of histidine phosphorylated protein in the lysate of microbial cells.

Key words: Hapten, Phosphohistidine antibody, Specificity, Avidity

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