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Bradford Protein Assay
Published: Mar 20, 2011 DOI: 10.21769/BioProtoc.45 Views: 516183
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Abstract
The Bradford protein assay is used to measure the concentration of total protein in a sample. The principle of this assay is that the binding of protein molecules to Coomassie dye under acidic conditions results in a color change from brown to blue. This method actually measures the presence of the basic amino acid residues, arginine, lysine and histidine, which contributes to formation of the protein-dye complex. Unlike the BCA assay, reducing agents (i.e., DTT and beta—mercaptoethanol) and metal chelators (i.e., EDTA, EGTA) at low concentration do not cause interference. However, the presence of SDS even at low concentrations can interfere with protein-dye binding. This technique was invented by Bradford (1976).
Materials and Reagents
- Bovine Serum Abumin (BSA) (Sigma-Aldrich)
- Coomassie Brilliant Blue G-250 (Sigma-Aldrich, catalog number: 27815 )
- Methanol
- Phosphoric acid (H3PO4)
- Bradford reagent (see Recipes)
Equipment
- Spectrophotometer (Tecan)
- Whatman #1 paper (Whatman)
Procedure
Category
Biochemistry > Protein > Quantification
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