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[目的]通过实验,比较不同染色方法对海马组织蛋白质双向凝胶电泳(2-dimensional gel electrophoresis,2-DE)分离效果的影响,为类似的海马组织蛋白分离研究提供参考。[方法]分离、制备大鼠海马组织总蛋白样品并进行蛋白定量,不同含量海马蛋白质经2-DE分离后分别采用改良考马斯亮蓝染色、硝酸银染色法,中浓度上样量的的凝胶经考马斯亮蓝染色后也用考马斯亮蓝-硝酸银进行复染,比较不同染色方法对2-DE分离效果的影响。[结果]不同染色方法随上样量增加,所见蛋白斑点增加,干扰物质也增加。在本研究条件下,700μg上样量采用硝酸银染色效果最佳,考马斯亮蓝则显示1800μg上样量效果更优。1200μg考马斯亮蓝-硝酸银复染的效果较单独考染效果好。[结论]硝酸银染色较改良考马斯亮蓝更敏感,凝胶所见蛋白质斑点更多,考马斯亮蓝-硝酸银复染效果较单独改良考马斯亮蓝好。
[Objective] The aim of the study was to compare the effects of different staining methods on the separation of 2-dimensional gel electrophoresis (2-DE) in hippocampal tissue and to provide a reference for the similar study on the separation of hippocampal tissue proteins. [Method] Total hippocampus tissue protein samples were isolated and quantified. The hippocampus proteins with different contents were separated by 2-DE, and then were separated by modified Coomassie brilliant blue staining, silver nitrate staining, After staining with Coomassie Brilliant Blue, Coomassie brilliant blue-silver nitrate was also used for counterstaining. The effects of different staining methods on 2-DE separation efficiency were compared. [Result] The different staining methods increased with the amount of sample, the spots of protein increased, and the interfering substances also increased. Under the conditions of this study, staining with silver nitrate at 700μg was the best, and Coomassie blue showed the best results at 1800μg. 1200μg Coomassie brilliant blue - silver nitrate double staining effect than the single test results. [Conclusion] Silver nitrate staining was more sensitive than modified Coomassie Brilliant Blue, and there were more protein spots on the gel. Coomassie Brilliant Blue - silver nitrate complex was better than Coomassie Brilliant Blue alone.