The method of Bradford is a spectroscopic method of analysis used to measure the concentration of the Protéine S in solution in water.

Principle

The method of Bradford is a colorimetric proportioning , based on the change wavelength of Absorbance, appearing by the change of the color of the Bleu of Coomassie after connection (complexation) with the Arginine and the hydrophobic residues of the Amino-acid present in proteins.

The anion form (dependant) of the dye is blue, and has a maximum Absorption spectrum estimated historically at 595 Nm. The cation shapes (free) of the dye are red and green, absorbent with 465-470 Nm. The change of absorptance is proportional to the quantity of bound dye, thus indicating the protein concentration in the sample.

Contrary to the other methods of measurement of proteins, the method of Bradford is less sensitive to the interferences by various agents present in the protein samples. It however is affected by the Détergent S, is modified by the pH, and also gives a positive test to water-soluble polyphenols of high molecular weight (tannins).

Disadvantages

• the method of Bradford is linear on a narrow interval, typically of 2 microg/ml with 120 microg/ml, which makes necessary of preliminary dilutions of the tissue sample before analysis.
• the amino-acids, the Peptide S and the Protéine S of low molecular weight (<3000 Da) are not detected by this method.
• Reacts positively to the Tanins in solution after crushing of vegetable fabrics. Any proportioning of soluble proteins total of a vegetable fabric by this method must be preceded by a fixing by these tannins by means of PVPP for example.

Another possible use

Proportioning of tannins in solution, by taking account however of the reaction with proteins in solution which is added again.

Other methods

Other methods of analysis and of measurement of proteins include

• the Spectroscopie by UV
• the Réaction of the biuret
• the Méthode of Lowry
• the bicinchoninic Méthode by acid

O-phthalaldehyde protein assay -->

Reference

• Bradford, Mr. Mr. (1976) has off Rapid and Sensitive Method for the Quantitation Microgram Quantities off Protein Utilizing the Principle off Protein-Dye Binding . Anal. Biochem. 72 : 248-254.
• Kilkowski, W.J., G.G. Large, 1999 Color reaction off hydrolyzable tannins with Bradford reagent, Coomassie brilliant blue . Phytochemistry. 51 : 363-366.

External bonds

• Bradford Assay Background
• Bradford Assay Protocol

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