Shaw Margaret M, Riederer Beat M
Institut de Biologie Cellulaire et de Morphologie, Université de Lausanne, Rue du Bugnon 9, 1005 Lausanne, Switzerland.
Proteomics. 2003 Aug;3(8):1408-17. doi: 10.1002/pmic.200300471.
The choice of sample preparation protocol is a critical influential factor for isoelectric focusing which in turn affects the two-dimensional gel result in terms of quality and protein species distribution. The optimal protocol varies depending on the nature of the sample for analysis and the properties of the constituent protein species (hydrophobicity, tendency to form aggregates, copy number) intended for resolution. This review explains the standard sample buffer constituents and illustrates a series of protocols for processing diverse samples for two-dimensional gel electrophoresis, including hydrophobic membrane proteins. Current methods for concentrating lower abundance proteins, by removal of high abundance proteins, are also outlined. Finally, since protein staining is becoming increasingly incorporated into the sample preparation procedure, we describe the principles and applications of current (and future) pre-electrophoretic labelling methods.
样品制备方案的选择是等电聚焦的一个关键影响因素，而这反过来又会在质量和蛋白质种类分布方面影响二维凝胶的结果。最佳方案会因待分析样品的性质以及要分离的组成蛋白质种类的特性（疏水性、形成聚集体的倾向、拷贝数）而有所不同。本综述解释了标准样品缓冲液的成分，并阐述了一系列用于处理二维凝胶电泳不同样品（包括疏水膜蛋白）的方案。还概述了当前通过去除高丰度蛋白质来浓缩低丰度蛋白质的方法。最后，由于蛋白质染色越来越多地被纳入样品制备过程，我们描述了当前（以及未来）预电泳标记方法的原理和应用。
