一种新的双向电泳蛋白样品制备方法

A New Preparation Method for the Protein Sample in Two-dimensional Electrophoresis

目的探讨双向凝胶电泳(2-DE)样品裂解液对蛋白质分离和双向凝胶电泳结果的影响。方法以MOLT-4细胞为例,采用3种溶解性能不同的裂解液提取细胞中的蛋白质组,并分别进行双向聚丙烯酰胺凝胶电泳。结果通过对2-D PAGE图谱的比较分析,发现采用8 mol/L Urea,2 mol/L Thiourea,4%CHAPS,1%NP-40,4%TritonX-100,65 mmol/L DTT,0.5 mmol/L PMSF,0.5%pharmalyte 3-10,能获得质量较高的2-DE图谱。结论采用高浓度脲、三去污裂解液有利于获得优质的蛋白质样品,从而提高蛋白质提取率和双向电泳分辨率等。

Objective To investigate the effects of the sample lysis solution on protein separation and results about two-dimensional electrophoresis in two-dimensional gel electrophoresis. Methods Three different resolvable solutions were adopted to take out the proteins of MOLT-4 cells as the example, and then the proteins were separated by two dimensional polyacrylam- ide gel electrophoresis. Results By using 8 mol/L Urea, 2 mol/L Thiourea, 4 % CHAPS, 1 % NP-40, 4 % TritonX-100, 65 mmol/L DTT, 0.5 mmol/L PMSF, 0.5 % pharrnalyte 3-10 much better quality map of 2-DE could be obtained. Conclusion Using lysis solution containing high concentration urea and three different detergents can obtain more quality samples, then increase the extraction rate of proteins and resolution about two-dimensional electrophoresis.