BCA Protein Assay

The BCA Protein Assay, also known as the Smith Assay, is a highly sensitive colorimetric assay that is compatible with detergent solubilized protein solutions and primarily relies on two reactions. Firstly, the peptide bonds in the protein sample reduce Cu²⁺ ions, in a temperature dependent reaction, from the copper solution to Cu⁺. The amount of Cu²⁺ reduced is proportional to the amount of protein present in the solution. Next, two molecules of bicinchoninic acid (BCA) chelate with each Cu⁺ ion, forming a purple-colored product that strongly absorbs light at a wavelength of 562 nm that is linear for increasing protein concentrations between the range of 0.02-2mg/ml. The amount of protein present in a solution can be quantified by measuring the absorption spectra and comparing with protein solutions with known concentrations. The Bicinchoninic Acid (BCA) Protein Assay is suitable for quantifying protein solutions in 1ml assays or in micro-wells.