The Bradford Assay: Protein Quantification

Part of the series: Biochemical Tools & Techniques

Biochemists often need to quantify protein concentrations in solutions. However, because of the diverse nature and composition of proteins, a single technique may not be the most accurate or efficient in all cases.

Marion M. Bradford, in 1976, developed what is now known as the Bradford Assay. It is quick and accurate in determining the concentration of proteins in a solution.

In an acidic solution, Coomassie Brilliant Blue R-250 dye (Figure 1) binds to proteins, and a shift in wavelength of maximum absorption from 465 nm to 595 nm. The clever aspect of this assay is that the absorption at 595 nm is directly proportional to the protein concentration. Using a calibration curve, a biochemist can measure the absorption and determine the protein concentration in an unknown solution. The sensitivity of the Bradford Assay (1 μg) is higher than other techniques, including the Biuret (1-20 mg) and Lowry (5 μg) assays.

However, the Bradford Assay is limited by solutions that can cause interferences. SDS (sodium dodecyl sulfate), a common detergent used in gel electrophoresis, can interfere with the binding of Coomassie Brilliant Blue R-250 dye to proteins, leading to an underestimated protein concentration.

Despite this limitation, the Bradford Assay is a quick and reliable technique used by biochemists today.

Figure 1. Coomassie Brilliant Blue R-250