Before #christmas we already showed you one assay to #quantify proteins in solution 🧪. Today, we present the #BCA assay which is used for the same purpose. The bicinchoninic acid (BCA) assay is used to determine total protein #concentration in a solution. The solution changes color from #green to purple in proportion to the #protein concentration and thereby the quantity can be determined #colorimetrically.
The BCA #assay relies on the reduction of Cu2+ to Cu+ by the protein in solution in an #alkaline medium. This is followed by colorimetric detection of the Cu+ by BCA.
The protein forms a colored chelate complex with #copper in an alkaline solution coloring the solution blue 💙. Then, BCA reacts with reduced Cu+ by forming an intensely purple chelation complex of two BCA #molecules with one Cu+ 🤩. This complex has a strong linear absorbance at 562 nm with increasing protein concentrations. Cysteines, tyrosines and tryptophans incluence the reaction but unlike assays with Coomassie like the #Bradford assay from last week, the #peptide backbone itself also contributes to color formation. This minimizes amino acid sequence dependent variability 🕵️.
Have a nice Thursday 🤗