The Bradford protein assay is based on the visual phenomena that occurs when the UV light absorbance maximum of an acidic solution of Coomassie Brilliant Blue G-250 shifts from 465 nm to 595 nm when it is bound to protein.
Hydrophobic and ionic interactions can both stabilize the anionic forms of the Bradford Coomassie dye yielding a noticeable visible color change.
Research indicates that the free dye can exist in four different ionic forms. Of the three charged forms of the dye, the more cationic red and green forms have absorbance maxima at 470 nm and 650 nm, respectively. In contrast, the more anionic blue form of the dye, which binds to protein, has an absorbance maximum at 590 nm. For more information see Bradford Assay
Also the color change is seen in the linear range of the Bradford assay from about 5-25 mcg/mL, and therefore the more protein that is bound the more Coomassie binds.
Bradford Assay Method The bradford dye-binding assay is a colorimetric assay for measuring total protein concentration. It involves the binding of Coomassie Brilliant blue to protein. There is no interference from cations nor from carbohydrates such as sucrose.
Detergents such as sodium dodecyl sulfate and triton x-100 can interfere with the assay, as well as strongly alkaline solutions.
Includes a general overview of the procedure and preparation of the standards in the protocol.
Bradford Protein Assay A detailed and well thought out Bradford protein assay page using a spectrophotometer. Includes information such as the Bradford assay is very fast and uses about the same amount of protein as the Lowry assay, comments, procedural steps, equipment used and more.
Bradford Protein Assay Spectrophotometry Bradford Protein Assay Spectrophotometry. Includes spectrophotometry information and the Bradford protein assay: A spectrophotometer or colorimeter makes use of the transmission of light through a solution to determine the concentration of a solute within the solution. A spectrophtometer differs from a colorimeter in the manner in which light is separated into its component wavelengths. A spectrophotometer uses a prism to separate light and a colorimeter uses filters.
Bradford Protein Concentration Assay Includes Abbreviations, Background, and Procedure steps using BSA. The Bradford protein assay (1) is one of several simple methods commonly used to determine the total protein concentration of a sample. The method is based on the proportional binding of the dye Coomassie to proteins. The assay is colorimetric; as the protein concentration increases, the color of the test sample becomes darker. Coomassie absorbs at 595 nm.