肝癌细胞线粒体的纯化及双向凝胶电泳分析

IsoIation and 2-DE analysis of mitochondria of cultured hepatoma cells

目的:建立纯化肝癌细胞线粒体的方法,通过双向电泳及酶活力测定确认纯化效果。方法:使用蔗糖密度梯离心法分离纯化线粒体,双向凝胶电泳分析纯化过程中样品蛋白质图谱的变化。结果:通过标志酶活力的监测,梯度离心后线粒体纯提高近12倍,线粒体蛋白质在双向凝胶电泳中得到很好的显现。结论:使用蔗糖密度梯度心法可有效地分离纯化线粒体。

Objective:To make the isolation and 2-DE analysis of mitochondria,in order to prepare the following proteomic study on mitochondria of hepatoma-cell. Methods:Mitochondria was separated from cell homogenate by means of density gradient centrifugation,and 2-DE was conducted to examine the protein profile of mitochondrial preparations.Results:The increase in purity of mitochondria was found to be 12 times by density gradient centrifugation.Mitochondrial proteins were displayed well in the 2-DE pattern after purification.Conclusion:The isolation procedure is practicable,which provides a basis for the following proteomic study on mitochondria.