小鼠神经组织线粒体提取方法研究

Research of Methods for Isolation of Mitochondrion from Nervous Tissue in Mouse

采用两次梯度离心的方法提取原代皮层神经元、皮层和海马组织线粒体,通过健那绿B染色、Western blot方法检测所提取线粒体的纯度.健那绿B染色结果显示,在600g离心力的情况下,经过第一次梯度离心,第二次梯度离心在1 000g离心力的情况下进行,所提取线粒体结构最为完整,纯度最好,几乎不含细胞碎片;而在3 000g离心力情况下,线粒体纯度最低,且含有较多的细胞碎片.Western blot方法检测结果显示,1 000g离心力情况下线粒体的纯度最高.以上研究结果表明,采用梯度离心法提取神经组织线粒体时,第二次梯度离心的离心力越低提取的线粒体纯度越高.

Mitochond centrifugations. The ria of primary cortical neurons, cortex and hippoeampus were isolated by two gradient purity of mitochondrion was detected by Janus green B staining and Western blotting. The mitochondrial structure and the purity are the best, without cell debris, by the second centrifugal force after the first centrifugal force with 600 g, however, the mitochondria contain more cell debris under the force of 3 000 g. The results of Western blot also show that the purity of mitochondria is the highest of 1 000 g centrifugal centrifugal force. This study shows that the lower the centrifugal force in secondly centrifugation, the higher is the obtained mitochondrial purity when the mitochondria are isolated by gradient centrifugation in neural tissue.