UNIT 10.4 Two-Dimensional Gel Electrophoresis

  1. Lonnie D. Adams1,
  2. Sean R. Gallagher2

Published Online: 1 SEP 2004

DOI: 10.1002/0471142727.mb1004s67

Current Protocols in Molecular Biology

Current Protocols in Molecular Biology

How to Cite

Adams, L. D. and Gallagher, S. R. 2004. Two-Dimensional Gel Electrophoresis. Current Protocols in Molecular Biology. 67:II:10.4:10.4.1–10.4.23.

Author Information

  1. 1

    The Upjohn Company, Kalamazoo, Michigan

  2. 2

    UVP, Inc., Upland, California

Publication History

  1. Published Online: 1 SEP 2004
  2. Published Print: JUL 2004


Two-dimensional gel electrophoresis is the combination of two high-resolution electrophoretic procedures (isoelectric focusing and SDS-polyacrylamide gel electrophoresis) to provide much greater resolution than either procedure alone. In the first-dimension gel, solubilized proteins are separated according to their isoelectric point (pI) by isoelectric focusing. This gel is then applied to the top of an SDS-slab gel and electrophoresed. The proteins in the first-dimension gel migrate into the second-dimension gel where they are separated on the basis of their molecular weight. The basic protocols in this unit are based on the type of equipment originally described by O'Farrell in 1975. For very basic or very acidic proteins, two alternate protocols are provided. A third alternate protocol describes how two-dimensional electrophoresis can be performed using a minigel system. Protein sample preparation is presented in the support protocol.