BRADFORD PROTEIN ASSAY

The 'Bradford Protein Assay' is a spectroscopic analytical procedure used to measure the concentration of protein in a solution.

Contents

Principle

Disadvantages

Alternative assays

Reference

External links

Principle

The Bradford assay, a colorimetric protein assay, is based on an absorbance shift in the dye Coomassie when bound to arginine and hydrophobic amino acid residues present in protein.
The (bound) form of the dye is blue and has an absorption spectrum maximum historically held to be at 595 nm. The anionic (unbound) forms are green and red. The increase of absorbance at 595 nm is proportional to the amount of bound dye, and thus to the amount (concentration) of protein present in the sample.
Unlike other protein assays, the Bradford protein assay is less susceptible to interference by various chemicals that may be present in protein samples. An exception of note is elevated concentrations of detergent.

Disadvantages

The Bradford assay is linear over a short range, typically from 2ug/ml to 120ug/ml, often making dilutions of a tissue sample necessary before analysis.
Because the Bradford assay essentially measures the amount of arginine and hydrophobic amino acid residues, the amino acid composition can alter the concentration-absorbance curve depending on the percentage of arginine or hydrophobic amino acids in each protein. It is therefore necessary to use a standard whose protein closely matches the measured protein in composition.

Alternative assays

Alternative protein assays include

★ UV spectroscopy

★ Biuret protein assay

★ Lowry protein assay

★ Bicinchoninic acid protein assay

★ Amido black protein assay

★ o-phthalaldehyde protein assay
The assay kit provided by SIGMA is linear for concentrations up to 1.4mg/ml

Reference

★ Bradford, M. M. (1976) ''A Rapid and Sensitive Method for the Quantitation of Microgram Quantities of Protein Utilizing the Principle of Protein-Dye Binding''. Anal. Biochem. '72':248-254.

External links

★ Bradford Assay Background

★ Bradford Assay Protocol