优化硝酸银染色在心肌线粒体蛋白双向电泳后凝胶染色中的应用

Application of Optimized Silver Staining in the Gel Dyeing of Myocardial Mitochondria Protein after Two-Dimensional Electrophoresis

蛋白组学研究中双向电泳(2-DE)后的SDS-PAGE凝胶染色存在灵敏度和质谱兼容性问题。线粒体中存在大量微量蛋白,对于这类亚细胞器蛋白质2-DE后的凝胶染色,这两种特性显得尤为重要。经典的硝酸银染色虽灵敏度高,但之后的质谱鉴定兼容性欠佳。因此,该研究通过优化硝酸银染色方法观察大鼠心肌线粒体蛋白质2-DE后的凝胶染色效果,挖取部分蛋白质斑点酶解后行基质辅助激光解吸电离飞行时间质谱(MALDI-TOF-MS)鉴定,验证优化硝酸银染色与质谱鉴定的兼容性。结果表明,优化后的硝酸银染色方法灵敏度高,与MALDI-TOF-MS兼容性好,是一种线粒体蛋白质2-DE后染色的理想方法。

There are the problems of the sensitivity and mass-spectrum compatibility for staining after twodimensional electrophoresis（2-DE） in proteomics. There are a lot of micro-proteins in mitochondria. These two properties are particularly important for the gel staining after 2-DE. Although the classic silver staining has a high sensitivity, the compatibility of mass spectrometry identification is low. Therefore, we optimized the silver staining for rat myocardial mitochondrial proteomics. We observed the image of gels and extracted some protein spots that were identified by matrix-assisted laser desorption ionization time-of-flight mass spectrometry（MALDI-TOFMS） to verify the compatibility of mass spectrometry. The experimental results showed that the optimized method of staining has high sensitivity and good compatibility with mass spectrometry. The optimized silver staining is an ideal method in myocardial mitochondrial proteomics.