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Electrophoresis of Proteins and Protein-Protein Complexes in Native Polyacrylamide Gels Using a Horizontal Gel Apparatus

Authors
Journal
Analytical Biochemistry
0003-2697
Publisher
Elsevier
Publication Date
Volume
223
Issue
1
Identifiers
DOI: 10.1006/abio.1994.1552
Disciplines
  • Biology

Abstract

Abstract Electrophoresis of proteins and protein-protein complexes in polyacrylamide gels under native conditions using a horizontal gel apparatus is described. The advantage of this system is that it permits the detection of both negatively and positively charged proteins as well as protein-protein complexes in the same gel. During electrophoresis, a continuous gel sandwiched between two glass plates is placed horizontally on the platform and submerged in a reservoir buffer. The sample wells are made along the center of the gel, allowing positively and negatively charged proteins to migrate toward the cathode and anode, respectively. Several proteins with varying molecular weights and isoelectric point (p I) values and pairs of proteins capable of forming protein-protein complexes were chosen as model systems to illustrate the methodology. The effects of several parameters on the performance of the gel system including protein molecular weight, p I, and gel concentration were also examined and the results obtained by this method are comparable to those obtained by the vertical system. Following electrophoresis, both negatively and positively charged proteins as well as protein-protein complexes can be transferred by electroblotting onto polyvinylidene difluoride membranes for further analyses.

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