Haynes:Bradford

Bradford Assay
by Karmella Haynes, 2012

Principle: The dye in the Bradford reagent turns from brown to blue in the presence of protein. The absorbance f blue color is proportional to the protein concentration. See http://en.wikipedia.org/wiki/Bradford_protein_assay

1. Label enough 1.5 mL eppendorf tubes for one blank (1) , five standard samples (2-6), and all of your unknown samples (7-n).
2. Add 500 μL Bradford Reagent to each tube. You will add protein to these later, and ignore the negligible change caused by additional protein volume.
3. Add a BSA standard protein solution* to tubes 2 (1μg BSA), 3 (2μg BSA), 4 (4μg BSA), 5 (8μg BSA), and 6 (16μg BSA). (*Note, use the appropriate volume based on the concentration of your stock BSA).
4. Add 5 μL of unknown to each remaining tube. Keep track of your samples with good labeling.
5. Transfer 200 μL of the blank (tube one) into the first del in a clear 96-well flat-bottom plate.