title: Horizontal comparative fluorescence two-dimensional gel electrophoresis for improved spot coordinate detection.
abstract: Vertical comparative 2D fluorescence gel electrophoresis (CoFGE) has recently been shown to increase the reproducibility of coordinate assignment for protein spots, in particular in singular experiments, which cannot be investigated using DIGE. The method applies a standardized marker grid formed by a set of purified proteins to the sample proteome in a conglomerate of 1DE, 2DE, and DIGE. Here, improvements are demonstrated by transferring CoFGE to horizontal 2DE. These include the elimination of the protein modification by residual acrylamide monomer unavoidable in vertical CoFGE, reduced buffer volumes, and highly efficient laboratory procedures. Spot patterns are well defined and can be easily analyzed using commercially available warping algorithms. With horizontal CoFGE also a correction for changes in pI was introduced using a third fluorescent dye. Horizontal CoFGE holds high promises in comparative proteomics.
authors.affiliation: Integrated Functional Genomics, Interdisciplinary Center for Clinical Research, University of Münster, Germany.
authors.forename: Marina
authors.initials: M
authors.lastname: Hanneken
authors.forename: Simone
authors.initials: S
authors.lastname: König
mesh_terms: D015180:Electrophoresis, Gel, Two-Dimensional / Q000295:instrumentation*; D004867:Equipment Design; D004926:Escherichia coli / Q000737:chemistry*; D029968:Escherichia coli Proteins / Q000302:isolation & purification*; D005453:Fluorescence; D005456:Fluorescent Dyes / Q000032:analysis; D040901:Proteomics / Q000295:instrumentation; D013194:Staining and Labeling
chemical_list: D029968:Escherichia coli Proteins; D005456:Fluorescent Dyes
keywords: G-Dyes; Horizontal 2DE; Horizontal comparative fluorescence gel electrophoresis; Protein coordinates; Protein grid