高等学校化学学报

高等学校化学学报

• 研究论文 • 上一篇    下一篇

β2-微球蛋白连续表位的免疫亲和质谱研究

黎根1,2, 刘宁1, 刘志强1, 刘淑莹1   

    1. 中国科学院长春应用化学研究所, 长春质谱中心, 长春130022;
    2. 中国科学院研究生院, 北京100049
  • 收稿日期:2005-12-05 修回日期:1900-01-01 出版日期:2007-01-10 发布日期:2007-01-10
  • 通讯作者: 刘淑莹

Characterization of Linear Epitope of the β2-Microglobulin via Combination of MALDI-TOF-MS with Immunoaffinity

LI Gen1,2, LIU Ning1, LIU Zhi-Qiang1, LIU Shu-Ying1   

    1. Changchun Institute of Applied Chemistry, Chinese Academy of Sciences, Changchun Center of Mass Spectrometry, Changchun 130022, China;
    2. Graduate School of the Chinese Academy of Sciences, Beijing 100049, China
  • Received:2005-12-05 Revised:1900-01-01 Online:2007-01-10 Published:2007-01-10
  • Contact: LIU Shu-Ying

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摘要： 采用Endoproteinase Glu-C, Lys-C和Trypsin 3种蛋白酶分别水解β2-微球蛋白, 产生一系列肽段, 利用固定在琼脂糖珠上的单克隆抗体与其发生免疫亲和反应. 利用基质辅助激光解吸电离飞行时间质谱(MALDI-TOF-MS)技术, 对抗原决定簇肽段-抗体复合物进行系统研究, 结果表明, 与抗体结合部位即连续表位的位点为肽段(59~69)(DWSFYLLYYTE). 该研究方法简便、准确, 可用来对其它抗原连续表位的快速测定.

关键词: 基质辅助激光解吸电离飞行时间质谱, 免疫亲和, β2-微球蛋白, 连续表位, 单克隆抗体

Abstract: Matrix-assisted laser desorption ionization time-of-flight mass spectrometry(MALDI-TOF-MS), in combination with immunoaffinity provided a powerful tool for determining epitope(antigenic determinant) in protein. The linear epitope of the β2-microglobulin was characterized in the paper. The method as follows: at first β2-microglobulin was digested by a proteolytic enzyme to produce an appropriate set of peptide fragments, then peptide fragments containing the linear epitope were selected and separated from the pool of peptide fragments by immunoprecipitation with the monoclonal antibody. The agarose beads were collected carefully after the reaction. Unbound peptides would be washed away, while the peptides containing the epitope would remain bound to the immobilized antibody after the beads were washed several times with appropriate buffer. At last the masses of the bound peptides were identified directly by MALDI-TOF MS. Using Endoproteinase Glu-C Endoproteinase Lys-C and Trypsin in the experiment, the linear epitope of β2-microglobulin was located within peptide fragment 59—69, that is, DWSFYLLYYTE.

Key words: MALDI-TOF-MS, Immunoaffinity, β2-Microglobulin, Linear epitope, Monoclonal antibody

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