Bradford test for protein

The Bradford assay is a very good, and simple, method of detecting microgram quantities of protein.

However the test is specific for certain amino acids, principally arginine, so not all proteins give the same reaction. For example albumin, casein and gelatin all give different responses. Gelatin has a very weak response to Bradford reagent since the protein, which is partially hydrolysed collagen, contains very few of the amino acids to which the reagent is sensitive. The standard usually employed is bovine serum albumin, (BSA). This is relatively expensive. We have used powdered egg white from the home-baking section of our local supermarket as the standard with results comparable to BSA.

The reagent contains Coomassie Blue dye which is light brown in the reagent but blue when bound to protein.


eye-protection The stock solution contains phosphoric acid (50%). corrosive
gloves Diluted for use the reagent is irritant.
The dye will stain skin and clothes.
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Bradford reagent

    Dissolve 50mg Coomassie Blue in 20cm3 methanol

    Add this to 60cm3 phosphoric acid

    Make up to 100cm3 with water

    Label the stock solution 'CORROSIVE'
    Dilute with 4 volumes of water for use

Add 2.5cm3 of the reagent, (stock solution diluted 1+4 with water), to 0.25cm3 of the sample solution.
Allow the mixture to stand for ten minutes then read the absorbance using red light. Ten minutes allows full development of the colour, longer intervals will not affect the result.

The sensitivity can be increased by adding 0.5cm3 of undiluted stock solution to 2cm3 of the solution to be tested. The method will detect concentrations down to about 5µg per cm3.

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    Egg albumin (powdered egg white) was the standard used for these results. Bradford graph

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