Quantitative determination of Coomassie R bound to proteins in polyacrylamide gel. A facilitated method for multi-spot analysis of electrograms.
Dimethylsulfoxide (DMSO) was found to be an efficient solvent for extraction of Coomassie Blue R 250 (Coomassie R) from stained proteins on polyacrylamide gels. Kinetic measurements show that the extraction of the dye from a 2-D gel reached equilibrium in 48 h. Staining of E. coli ribosomal proteins by Coomassie R dissolved in trichloroacetic acid exhibited two types of dye-protein complexes, the majority of them yield a blue-purple colour, while the rest are stained with a light-blue tone and fluorescent appearance as well. The absorbance spectra of the complexes in the gel matrix differ significantly from each other. However, the DMSO-extracted Coomassie show identical absorbance profiles with lambda max at 602 nm, thus the amount of the bound dye can easily be measured spectrophotometrically.
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D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000032:analysis; D007700:Kinetics; D007930:Leucine / Q000032:analysis; D011506:Proteins / Q000032:analysis*; D012270:Ribosomes / Q000032:analysis; D012394:Rosaniline Dyes / Q000032:analysis*; D012997:Solvents; D013194:Staining and Labeling
D011506:Proteins; D012394:Rosaniline Dyes; D012997:Solvents; C024757:Coomassie brilliant blue R; D007930:Leucine