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. 2021:2228:53-62.
doi: 10.1007/978-1-0716-1024-4_5.

Proteome Analysis with Classical 2D-PAGE

Caroline May  1   2 , Frederic Brosseron  1 , Kathy Pfeiffer  1   2 , Kristin Fuchs  1   2   3 , Helmut E Meyer  1 , Barbara Sitek  1   2   3 , Katrin Marcus  4   5
Affiliations

Proteome Analysis with Classical 2D-PAGE

Caroline May et al. Methods Mol Biol. 2021.

Abstract

Two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) is based on the combination of two orthogonal separation techniques. In the first dimension, proteins are separated by their isoelectric point, a technique known as isoelectric focusing (IEF). There are two important variants of IEF, which are carrier-ampholine (CA)-based IEF and immobilized pH-gradient (IPG)-based IEF. In the second dimension, proteins are further separated by their electrophoretic mobility using SDS-PAGE. Finally, proteins can be visualized and quantified by different staining procedures such as Coomassie, silver staining, or fluorescence labeling. This article gives detailed protocols for 2D-PAGE, using both CA- and IPG-based separation in the first dimension.

Keywords: Carrier ampholyte system (CA); Immobilized pH gradient (IPG); Isoelectric focusing (IEF); Sodium dodecyl sulfate (SDS); Two-dimensional polyacrylamide gel Electrophoresis (2D-PAGE).

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