Staining of Preparative 2-D Gels: Coomassie Blue and Imidazole-Zinc Negative Staining
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The identification of proteins using preparative gel electrophoresis and mass spectrometry requires reversible staining of relatively thick (1–1.5 mm) polyacrylamide gels. We have found that staining with colloidal Coomassie brilliant blue G-250 or negative staining with imidazole-zinc yields high-resolution stains (Fig. 1 ) that are compatible with subsequent mass spectrometric analysis ( see Notes 1 and 5 ).
Fig. 1. Preparative 2-D gels stained by the imidazole-zinc negative stain (left) and colloidal Coomassie blue G-250 stain (right). Gels were scanned by a Computing Densitometer (Molecular Dynamics, CA). Top: 1 mg of protein from ME-180 cervical carcinoma cells was separated by carrier ampholyte IEF and 11% SDS-PAGE, and then stained using the imidazole-zinc stain. Bottom: 1 mg of protein from A375 human melanoma cells was separated by carrier ampholyte IEF and 11% SDS-PAGE, and then stained using the colloidal Coomassie blue G-250.
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