几种双向凝胶电泳蛋白质检测方法的比较.docVIP

几种双向凝胶电泳蛋白质检测方法的比较 nbsp;nbsp;nbsp;nbsp;nbsp;nbsp;nbsp;nbsp;nbsp; 作者：秦慧，刘霆，柳斌，宋鑫，黄欣，杨金亮，赵霞，魏于全nbsp;nbsp;nbsp; 【摘要】nbsp; 本研究对蛋白质染色法进行比较和分析，以求获得满足蛋白质组研究所需的适宜的蛋白质检测法。实验采用2-D电泳分离急性早幼粒白血病细胞株NB4的全蛋白及1-D电泳分离蛋白质分子量标准物，从蛋白质检测的灵敏性、质谱兼容性、操作的简便性等角度，比较了传统考马斯亮蓝染色法、胶体考马斯亮蓝染色法、改良考马斯亮蓝染色法和银染法4种蛋白质着色法对凝胶分离蛋白质的检测影响，探讨了影响蛋白质染色与鉴定的影响因素。结果显示，银染检测灵敏度最高，但质谱鉴定相容性差，鉴定率近10%；改良考马斯亮蓝染色法检测灵敏度也高，可达4 ng/spot，质谱鉴定率可接近55%。结论： 改良考马斯亮蓝染色法的蛋白质检测灵敏度高，与质谱兼容好，能满足蛋白质组研究需要。 【关键词】nbsp; 蛋白质组 　　Abstractnbsp;nbsp;nbsp; The aim of this study was to compare and analyze protein staining in order to select the optimal staining method for proteomic research. Proteins from acute promyelocytic leukemia cell line NB4 and protein molecular weight marker were separated respectively by 2-D or 1-D electrophoresis and detected respectively by the typical Coomassie brilliant blue，the colloidal Coomassie brilliant blue，the modified Coomassie brilliant blue and the silver staining protocols. The protein detection sensitivity，compatibility with mass spectrometry (MS) and facility of the four staining protocols were compared. The results indicated that the silver staining exhibited the highest sensitivity and MS showed the lowest compatibilitynbsp; 10% ofnbsp; protein identification rate.nbsp; The detection sensitivity of the modified Coomassie brilliant blue staining was superior to that of other two Coomassie brilliant blue stainings，close tonbsp; but lower than the silver staining，however the compatibility with MS was better （protein identification rate about 55%）.nbsp;nbsp; It is concluded that the protein detection sensitivity of the modified Coomassie brilliant blue staining is high，and its compatibility with MS is better，thisnbsp; modified Coomassie brilliant blue staining is an optimal staining method for proteomic research. 　　Key wordsnbsp;nbsp; proteomics；two-dimensional gel electrophoresis；protein detection；Coomassie brilliant bluenbsp; staining；silver staining 　　蛋白质组研究要求有高分辨率的蛋白质分离