Two-Dimensional Polyacrylamide Gel Electrophoresis

Two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) separates proteins by their iso-electric points (IEPs) and electro-phoretic mobilities (1 ). In the first dimension, a sample is applied to an iso-electric focusing (IEF) acrylamide gel. Within this the con stituent proteins migrate through a pH gradient until they reach their IEPs. A protein’s IEP is the pH value where it has no net charge. This is as a result of the positively and negatively charged amino acids cancelling each other. Proteins in a sample thus become focused at their individual IEPs. By the end of the electro-focusing there will be essentially no movement of proteins. An alternative to iso-electric focus ing is nonequilibrium pH gradient electrophoresis (NEPHGE) (2 ).