UNIT 10.2A One-Dimensional SDS Gel Electrophoresis of Proteins

  1. Sean R. Gallagher

Published Online: 1 MAY 2001

DOI: 10.1002/0471142727.mb1002as47

Current Protocols in Molecular Biology

Current Protocols in Molecular Biology

How to Cite

Gallagher, S. R. 2001. One-Dimensional SDS Gel Electrophoresis of Proteins. Current Protocols in Molecular Biology. 10:II:10.2A.

Author Information

  1. Motorola Corporation, Tempe, Arizona

Publication History

  1. Published Online: 1 MAY 2001
  2. Published Print: JUL 1999

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Electrophoresis is used to separate complex mixtures of proteins, (e.g., from cells, subcellular fractions, column fractions, or immunoprecipitates) to investigate subunit compositions, and to verify homogeneity of protein samples. It can also serve to purify proteins for use in further applications. In polyacrylamide gel electrophoresis, proteins migrate in response to an electrical field through pores in the gel matrix; pore size decreases with higher acrylamide concentrations. The combination of gel pore size and protein charge, size, and shape determines the migration rate of the protein. In this unit, the standard Laemmli method is described for discontinuous gel electrophoresis under denaturing conditions, that is, in the presence of sodium dodecyl sulfate (SDS). Both full-size and minigel formats are detailed. Several alternate protocols are provided for specific applications. The first two alternate protocols cover electrophoresis of peptides and small proteins, separations that require modification of standard buffers: either a Tris-tricine buffer system, or modified Tris buffer in the absence of urea. Continuous SDS-PAGE is a simplified method in which the same buffer is used for both gel and electrode solutions and the stacking gel is omitted. Other protocols cover the preparation and electrophoresis of various types of gels: ultrathin gels, multiple single-concentration gels, gradient gels, multiple gradient gels, and multiple gradient minigels.