title: Isoelectric focusing and two-dimensional gel electrophoresis.
abstract: By far the highest resolution of all separation techniques for intact proteins in a single analytical run continues to be by the combination of isoelectric focusing (IEF) and SDS-PAGE, originally introduced by O'Farrell [(1975). J. Biol. Chem.250, 4007-4021]. This analytical platform has seen a number of significant advances and applications over the past 25years, including reproducibility using immobilized pH gradient (IPG) strips [Bjellqvist et al. (1982). J. Biochem. Biophys. Methods6, 317-339.], resolution in alkaline IEF using hydroxyethyldisulfide (HED) [Olsson et al. (2002). Proteomics2, 1630-1632], and quantification for differential expression proteomics on intact proteins on a global scale [DIGE; Unlu et al. (1997). Electrophoresis18, 2071-2077]. These major improvements will be highlighted in this chapter alongside the principle and theory of 2D gel electrophoresis, as well as detailed methods for general 2D gel electrophoresis best use protocols.
authors.affiliation: Mass Spectrometry Research Center, Vanderbilt University, Nashville, Tennessee, USA.
authors.forename: David B
authors.initials: DB
authors.lastname: Friedman
authors.forename: Sjouke
authors.initials: S
authors.lastname: Hoving
authors.forename: Reiner
authors.initials: R
authors.lastname: Westermeier
mesh_terms: D000818:Animals; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007202:Indicators and Reagents / Q000737:chemistry / Q000600:supply & distribution; D007525:Isoelectric Focusing / Q000379:methods; D008954:Models, Biological; D011506:Proteins / Q000032:analysis / Q000737:chemistry
chemical_list: D007202:Indicators and Reagents; D011506:Proteins