Protein Concentration Measurement
 
 
 

BioRad Protein Assay and the Bradford method


BioRad protein assay is based on the Bradford method and uses an acidic dye (Coomassie Brilliant Blue G-250 dye) which changes color depending on the amount of protein in a solution; the dye changes its absorbance maximum from 465 to 595nm when binding to protein. Spectrophotometric measurement is performed at 595nm for the Bradford method.





Dilute Bradford reagent 1:5
Prepare BSA protein standards using the relevant buffer used to extract your samples.
Suggested dilutions: 100; 50; 10; 5; 1; 0.5µg/µl

Dilute samples and protein standards in parallel: 1:100 in water (to reduce contribution of detergents, which will affect the measurement).
Don't forget to include the "blank" already at this stage.
Add 5µl of samples, standards and blank to 495µl diluted Bradford reagent.
Don't forget to include the "blank" already at this stage.

Incubate for 5min (but no more than one hour)

Measure absorbance and establish concentration curve using standards
Measure standards