Gelfi Cecilia, Capitanio Daniele
Dipartimento di Scienze Biomediche per la Salute, Università degli Studi di Milano, via f.lli Cervi, 93, 20090, Segrate, Milan, Italy.
U.O. Proteomica clinica, IRCCS Policlinico San Donato, 20097, San Donato, Milan, Italy.
Methods Mol Biol. 2018;1664:117-136. doi: 10.1007/978-1-4939-7268-5_11.
Two-dimensional difference gel electrophoresis (2-D DIGE) is an advanced and elegant gel electrophoretic analytical tool for comparative protein assessment. It is based on two-dimensional gel electrophoresis (2-DE) separation of fluorescently labeled protein extracts. The tagging procedures are designed to not interfere with the chemical properties of proteins with respect to their pI and electrophoretic mobility, once a proper labeling protocol is followed. The two-dye or three-dye systems can be adopted and their choice depends on specific applications. Furthermore, the use of an internal pooled standard makes 2-D DIGE a highly accurate quantitative method enabling multiple protein samples to be separated on the same two-dimensional gel. The image matching and cross-gel statistical analysis generates robust quantitative results making data validation by independent technologies successful.
二维差异凝胶电泳(2-D DIGE)是一种先进且出色的凝胶电泳分析工具,用于比较蛋白质评估。它基于对荧光标记蛋白质提取物进行二维凝胶电泳(2-DE)分离。一旦遵循适当的标记方案,标记程序设计为不干扰蛋白质关于其等电点(pI)和电泳迁移率的化学性质。可以采用双染料或三染料系统,其选择取决于具体应用。此外,使用内部混合标准使2-D DIGE成为一种高度准确的定量方法,能够在同一二维凝胶上分离多个蛋白质样品。图像匹配和跨凝胶统计分析产生可靠的定量结果,使得通过独立技术进行数据验证得以成功。