Zinc/Imidazole procedure for visualization of proteins in gels by negative staining

INTRODUCTIONThe zinc/imidazole staining procedure for visualizing proteins in acrylamide gels is based on differential salt binding. Because protein-bound salts (e.g., dodecyl sulfate or the heavy cation zinc) are chemically less active than free zinc ions in the gel, precipitation of an insoluble salt is much slower in those regions of the gel occupied by proteins than in the gel background where zinc dodecyl sulfate precipitates. The result is a "negative stain," with translucent proteins and an opaque gel background, due to zinc dodecyl sulfate precipitation. The sensitivity of the method has been markedly improved by altering the composition of the precipitated salt to a complex of zinc and imidazole. This protocol provides two methods: reverse stain using imidazole, SDS, and zinc, and double-staining using Coomassie blue stain followed by zinc/imidazole.