Microchip free flow isoelectric focusing with immobilized pH gradient on monolithic materials 基于固定化pH梯度(IPG)整体材料的芯片自由流等电聚焦电泳模式的构建
Methods : A 2-DE technique, including isoelectric focusing ( IEF ) with an immobilized pH gradient ( IPG ) and SDS-PAGE, was used. Sample preparation, rehydration, SDS-PAGE and other steps were established and optimized. 方法:采用固相pH梯度(IPG)等电聚焦(IEF)为第一向、SDS-PAGE垂直电泳为第二向的双向电泳技术,对样品的处理、水化、等电聚焦、SDS-PAGE等步骤进行了优化。
The resolution of 2-DE was increased after using the nonlinear pH3-10 or overlapping narrow immobilized pH gradient strips, and 11 differentially expressed protein spots were identified after analysis using the software. 采用了非线性pH3-10及重叠窄范围的固相pH梯度胶条,提高了2-DE图谱的分辨率;初步选择11个差异表达蛋白点。
Methods Healthy adult man bone marrow proteome of Han nationality was resuspended in two different solution systems. The extracted protein was separated by 2-DE, immobilized pH gradient ( IPG ) isoelectric focusing as the first dimension, then vertical SDS-PAGE as the second dimension. 方法人骨髓分别用A、B两种不同的溶解体系提取蛋白质,以固相pH梯度(IPG)等电聚焦为第一向,SDS-PAGE垂直电泳为第二向进行2-DE。
Conclusions Nonlinear pH3-10 and overlapping narrow immobilized pH gradient strips have higher separation efficiency for protein in 2-DE. The differentially expressed proteins separated will be useful for identifying metastasis-associated proteins of colorectal carcinoma and for establishing 2-DE database of the metastatic colorectal carcinoma cell lines. 结论非线性及重叠窄范围的固相pH梯度胶条可改善2-DE中蛋白分离的效果,初步分离的差异蛋白点为进一步鉴定大肠癌转移相关蛋白及建立大肠癌转移性细胞株的2-DE数据库奠定了基础。