Abstract,
Good control, efficiency, and reproducibility of protein extraction from cells and tissues are essential aspects for diverse biological and basic research applications. Effective and specific enzymatic digestion of proteins prior to mass spectrometry (MS) analysis is one of the fundamental techniques most commonly used in any proteomics laboratory. However, neither routine procedures for cell and tissue lysis nor for in-solution and in-gel protein digestion have been significantly altered in common practice for over a decade. Here we have tested and optimized several alternative techniques for preparing lysates of mammalian cells, as well as in-solution and in-gel enzymatic digestion compatible with downstream qualitative and quantitative MS-Based proteomics applications.
摘要,
从细胞和组织提取蛋白质的技术中, 良好的控制,效率和可重复性对于不同生物的分析和基础研究的应用是一个非常重要的观念。在质谱( MS )分析前, 有效和具体的蛋白酶解是一种蛋白质组学实验室中zui常用的基本技术。然而,无论是细胞和组织溶解的例行程序,或是凝胶蛋白质消解, 这十多年已被大大改进。基于蛋白质组学应用, 我们进行了几个哺乳动物细胞裂解的测试和优化, 以及在质谱( MS )的基础下进行in-solution and in-gel蛋白酶解的定性和定量分析。
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