免疫亲和质谱法研究β2-微球蛋白抗原表位characterization epitope of .pdf

2008 年第66 卷 化 学 学 报 Vol. 66, 2008 第13 期, 1572～1576 ACTA CHIMICA SINICA No. 13, 1572～1576 ·研究论文· 免疫亲和质谱法研究β -微球蛋白抗原表位 2 万翠红 黎 根 刘志强 刘淑莹* ( 中国科学院长春应用化学研究所 长春质谱中心 长春 130022) 摘要 采用免疫亲和分离与质谱分析相结合的方法, 对β2-微球蛋白抗原表位进行了系统研究. 完整的抗原分子和已固 定在载体(CNBr-activated Sepharose beads)上的单克隆抗体发生免疫亲和反应后, 用 Endoproteinase Glu-C, Trypsin, Aminopeptidase M 和carboxypeptidase Y 四种不同的蛋白酶依次酶解抗原分子, 并采用基质辅助激光解析电离飞行时间 质谱(MALDI-TOF-MS)技术对与抗体连接受保护而未发生水解的肽段进行了研究. 结果表明: β2-微球蛋白抗原表位位 于整个蛋白分子氨基酸序列的61～67 位, 即为SFYLLYY. 通过合成肽段的分析, 证明了SFYLLYY 即为抗原表位, 与 亲和质谱方法分析结果一致. 关键词 β2-微球蛋白; 抗原表位; 免疫亲和; 质谱 Characterization Epitope of the β -Microglobulin with Immunoaffinity 2 Mass Spectrometry WAN, Cui-Hong LI, Gen LIU, Zhi-Qiang LIU, Shu-Ying* (Changchun Center of Mass Spectrometry, Changchun Institute of Applied Chemistry, Chinese Academy of Sciences, Changchun 130022) Abstract The approach for epitope mapping is the application of immunoaffinity separation and matrix assisted laser desorption/ionization (MALDI) mass spectrometry (MS). The epitope of β2-microglobulin was identified with this approach. The antigen β2-microglobulin was coupled to a monoclonal antibody, which was covalently immobilized on a cyanogen bromide (CNBr)-activated Sepharose bead. Consecutive diges- tion of the antigen with endoproteinase Glu-C, trypsin, aminopeptidase M and carboxypeptidase Y resulted in