TWO DIMENSIONAL ELECTROPHORESIS (2DE) is one of the best experimental tools for the reliable separation of thousands of proteins in a single gel. 2DE consists of a tandem pair of electrophoretic separations:

In the first dimension, proteins are resolved in according to their isoelectric points (pIs) using immobilized pH gradient electrophoresis (IPGE), isoelectric focusing (IEF), or non-equilibrium pH gradient electrophoresis (NEPHGE). Under standard conditions of temperature and urea concentration, the observed focusing points of the great majority of proteins using IPGE (and to a lesser extent IEF) closely approximate the predicted isoelectric points calculated from the proteins' amino acid compositions.

In the second dimension, proteins are separated according to their approximate molecular weight using sodium dodecyl sulfate poly-acrylamide-electrophoresis (SDS-PAGE). This technique can provide molecular weight approximations (+/- 10%) for most proteins, with some dramatic exceptions.

Also see: from the SWISS-2DPAGE team at the ExPASy Molecular Biology Server or the Danish Center for Human Genome Research