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Bicinchoninic Acid Assay
The bicinchoninic acid assay (BCA), or Smith assay, is a method for measuring the total protein concentration of a solution. Generally serial dilutions of a protein standard, usually bovine serum albumin (BSA), are added to a multiwell plate in triplicate at known concentrations. Dilutions of the solutions to be tested are also added in triplicate. A solution of light green-colored cupric sulfate is added to each well and the plate is incubated at 37°C. At the higher temperature, the protein peptide bonds reduce the copper ions and bicinchoninic acid interacts with, or chelates the ions, resulting in a color change from light green to purple. The absorbance of 562 nm light by the purple, chelated copper ions is then measured. The absorbances of the samples are then compared to the BSA standards to determine protein concentration. Unlike many other protein quantification methods, BCA is detergent-compatible. BCA is a common lab technique with uses including standardizing protein concentrations prior to further analysis like gel electrophoresis and for measuring recombinant protein production. (Credit: Brooke Anderson-White)
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