High-Resolution, 2-D Protein Electrophoresis Using Nondedicated Equipment

Several formats—small, standard, and large—can be employed to separate proteins by two-dimensional (2-D) protein electrophoresis. The number of proteins in the sample as well as the degree of resolution needed determine the particular format used. The small format is preferred when few proteins (usually <30 proteins that differ to a moderate extent in size and charge) are to be separated. This format allows for a significantly shortened analysis time. The small gels also are easier to handle than larger ones. Large formats are useful for separating large numbers of proteins (usually >200), many of which may have similar molecular sizes or charges. Large formats provide the highest degree of separating power of all the formats, although some resolution with respect to spot intensity may be lost in some areas of the gel. The standard format is the most commonly employed format, because it provides a significantly higher degree of separating power than the small format and improved spot intensity over the large format. The gels also are much easier to handle than the more cumbersome and fragile large format gels.