Exercise 14: Protein Assay

The Coomassie Blue dye binding assay is a sensitive assay for determining protein concentration in samples and is based on the method originally developed by Bradford. The assay is reproducible, straightforward and easy to perform. Furthermore, the dye is compatible with various biochemical reagents, e.g. detergent solutions, buffers, chaotropic salts, preservatives such as sodium azide, some acids such as HCI etc. The proteins dissolved or mixed in any of these reagents can be measured. The assay is based on the principle of an absorbance shift resulting from an acidic Coomassie Brilliant Blue G-250 solution. When the dye solution is added to a protein sample interaction of the dye’s sulfonic groups with the protein results in a color change from brown to blue. This change can be measured at 595, 650 and 470 nm corresponding to anionic, neutral and cationic equilibrium states respectively for the