A Rapid and Sensitive Method for the Quantitation of Microgram Quantities of Protein Utilizing the Principle of Protein-Dye Binding

The Bradford Assay is well known for its ability to detect and quantify protein in solution with a great degree of sensitivity. During the assay, Coomassie Blue dye is allowed to bind to the protein for a carefully measured period of time, and then the samples are analyzed by visible spectrophotometry. Construction of a standard curve reveals an extinction coefficient that correlates absorbance of light at 595 nm with the concentration of protein in the test solution. For relatively pure protein solutions, the correlation is excellent and consistent however, presence of nucleic acids in a crude protein preparation has potential for interference. Interactions between Coomassie Blue Dye (G-250) and DNA from salmon testes during the Bradford Assay were characterized. It was found that Coomassie Blue G-250 in Bradford Assay reagent does interact with DNA at approximately one-fifteenth the rate of the interactions with standard bovine serum albumin.

Coomassie Brilliant Blue

Absorbance

Bradford protein assay

Bicinchoninic acid assay

Standard curve

Bovine serum albumin

Spectrophotometry

Molar absorptivity

10.1096/fasebj.21.5.a262-d

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The influence of sodium hyaluronate molecular weight on protein content according to Lowry and Coomassie blue assays

Carbohydrate Polymers (2008)

Claes Melander Kristoffer Tømmeraas

Coomassie Brilliant Blue

Bradford protein assay

Lowry protein assay

Molecular mass

Bicinchoninic acid assay

10.1016/j.carbpol.2008.04.001

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Citations (17)

Highly Sensitive Protein Concentration Assay over a Wide Range via Surface-Enhanced Raman Scattering of Coomassie Brilliant Blue

Analytical Chemistry (2010)

Xiao Han Yunfei Xie Bing Zhao Yukihiro Ozaki

In the Bradford protein assay, protein concentrations are determined by the absorbance at 595 nm due to the binding of Coomassie brilliant blue G-250 (CBBG) to proteins. In a protein−CBBG liquid mixture, surface-enhanced Raman scattering (SERS) is sensitive to the amount of unbound CBBG molecules adsorbed on silver surfaces, and the bound CBBG amount is directly related to the target protein concentration. Accordingly, a novel method for detecting total protein concentration in a solution has been developed based on SERS of unbound CBBG with an internal standard of silicon. Two obvious advantages of the proposed protein assay over conventional Bradford protein assay are its much wider linear concentration range (10−5−10−9 g/mL) and 200 times lower limit of detection (1 ng/mL), which demonstrates its great potential in rapid, highly sensitive concentration determination of high and low-abundance proteins.

Coomassie Brilliant Blue

Bradford protein assay

Absorbance

Bicinchoninic acid assay

10.1021/ac100596u

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Citations (65)

Bradford Assay for Determining Protein Concentration

Cold Spring Harbor Protocols (2020)

Clara L. Kielkopf William R. Bauer Ina L. Urbatsch

The Bradford assay is a quick and fairly sensitive method for measuring the concentrations of proteins. It is based on the shift in absorbance maximum of Coomassie Brilliant Blue G-250 dye from 465 to 595 nm following binding to denatured proteins in solution.

Coomassie Brilliant Blue

Absorbance

Bradford protein assay

Bicinchoninic acid assay

10.1101/pdb.prot102269

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Citations (338)

Coomassie blue protein dye-binding assays measure formation of an insoluble protein-dye complex

Analytical Biochemistry (1992)

T. C. Marshall Katherine M. Williams

Coomassie Brilliant Blue

Bradford protein assay