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This assay is based on the equilibrium between three forms of Coomassie Blue G dye. Under acidic conditions, proteins bind to the reagent, causing a spectral shift from reddish/brown to blue. Spectrophotometric measurement of the resulting level of blue is used to determine the concentration of protein solutions.
Protein + Coomassie Blue G => Red <=> Green <=> Blue-Protein (465 nm) (650 nm) (595 nm)
SPECIAL FEATURES
•Reducing agents used in protein samples do not interfere with this reagent.
•Fast, almost immediate, color development and ready for reading at 595 nm.
•Concentration range for protein assay from 1-1,500µg/ml.
•Microwell plate compatible.
•Easy to use.
•Inexpensive.
KIT CONTENTS
•Bradford Assay Reagent: For 500 Standard Assays, 1,000 Micro Assays and up to 5,000 microplate assays.
•Protein Standard Solution: 10 ml
Advantages and disadvantages using Coomassie-based protein assay:
The Bradford assay is a fast and easy method for protein assays. It does not need heating or special equipment. It is also compatible with most salts, solvents, buffers, reducing chemicals and chelating agents that are commonly found in protein samples.
The main disadvantage of Coomassie-based protein assays is their incompatibility with higher concentrations of surfactants, which are often used to solubilize membrane proteins. Generally, surfactant concentrations higher than 0.12% (such as SDS and Triton X-100), or 0.06% (such as Tween 20) in the sample, will cause precipitation of the reagent. If this is the case,the BCA Protein Assay(G1002)or Low Protein BCA Protein Assay(G1003)should be used.
REFERENCES
•Bradford, 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.
•Stoscheck, C. 1990, Quantification of Protein. Methods in Enzymology , 182:50-68.
•Lowry, O., et al.,1951, Protein measurement with the Folin phenol reagent. J. Biol.Chem.193:265-275.
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