Refinement of the Coomassie blue method of protein quantitation: A simple and linear spectrophotometric assay for ≤0.5 to 50 μg of protein
Abstract
The Coomassie brilliant blue G assay for proteins described by Bradford (1976) (Anal. Biochem.72, 248) was reexamined. It was found that the extinction coefficient of the dye-protein complex solution remained constant over the protein concentration range of 0.8 to 10 μg/ml of solution. This unchanging extinction coefficient, of protein/ml of solution, enhances both the sensitivity and versatility of the assay. Selection of a volume of dye-reagent (0.5 to 5.0 ml) which dilutes the protein sample to a final concentration of 0.8 to 10 μg/ml permits the application of Beer's Law for accurate determinations of ≤0.5 to 50 μg of protein. A combination of Bradford's study and the present one indicates that most common laboratory reagents and chemicals exert little or no influence on the A595 of the dye-reagent.
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