Crystal Violet as an Indicator Dye for Nonequilibrium pH Gradient Electrophoresis (NEpHGE)

Abstract

I found that a blue-colored dye, crystal violet, is a useful indicator for nonequilibrium pH gradient electrophoresis (NEpHGE). Unlike isoelectrofocusing, NEpHGE basically depends on the nonequilibrium gradient; therefore, the location of the proteins differs with acrylamide concentration or voltage used. To get reproducible results, colored marker which indicates the end of the electrophoresis might be very helpful. Crystal violet is an adequate dye for this purpose. This deeply colored, nontoxic material has a positive charge and migrated slightly faster than almost all proteins on NEpHGE. This dye affected neither the formation of pH gradient nor migration of proteins. Moreover, the ratio of migration (protein/dye) in the NEpHGE gels did not change under different conditions tested. Therefore, the addition of trace amounts of this dye gives a convenient endogenous color indicator to determine the end of the NEpHGE. Other positively charged dyes could be used as such an indicator, but some dyes (for example, methylene blue) lost their color by reduction during electrophoresis. Some dyes (for example, ethidium bromide) migrate too fast; therefore, they were not suitable indicators for NEpHGE.