大鼠皮层线粒体双向凝胶电泳体系的建立

Establishment of Two-Dimensional Gel Electrophoresis System for Rat Cortex Mitochondria

目的构建稳定的大鼠皮层线粒体双向凝胶电泳体系。方法 SD大鼠在规定的时间内快速断头取脑，冰上分离大脑皮层组织，两次密度梯度离心纯化线粒体。利用双向凝胶电泳技术改良等电聚焦的电压存3500V聚焦，总伏小时同定为14000Vh。以12％的SDS，PAGE进行第二向电泳．银染显色。结果得到了分辨率较高的皮层线粒体蛋白双向凝胶电泳图谱。结论利用改进的方法构建了高质量的双向电泳图谱，为各种疾病状态下皮层线粒体差异蛋白的研究提供了实验基础。

Objective To establish two-dimensional gel electrophoresis system for rat cortex mitochondria. Methods The adult male SD rats were sacrificed by cervical dislocation, the brain tissues quickly harvested and thecortex isolated on ice. The mitochondria were extracted and purified twice by density gradient centrifugation. Two-dimensional gel electrophoresis was performed. Immobilized pH gradient isoelectric focusing （IPG-IEF） was modified and carried out. The gels were focused at 3500 V for a total of 14000Vh and then subjected to SDS-PAGE and silver stai- ning. Results Electrophoregram of rat cerebral cortex mitochondria was obtained with high resolution by using the established two-dimensional gel eleetrophoresis system. Conclusion The two-dimensional gel electrophoresis system for rat cortex mitochondria was successfully established, which provides a theoretical basis and a technical support for research of cerebral cortex mitochondrion proteins in diseases.