Biuret is the method commonly used. In this method, the CO—NH groups of proteins form a purple complex with copper ions in an alkaline medium. As all proteins contain peptide bonds, this method is fairly specific. Biuret reagent is a mixture of alkaline copper sulphate and sodium potassium tartarate. The cupric ions form a CO—ordination complex with four —NH groups present in peptide bonds, giving an absorption maximum at 540–560 nm wavelength. Standard protein is also simultaneously prepared bovine serum albumin in the concentration 5 mg/ml and run along with protein sample including the Biuret reagent, and the optical density of each sample and standards are read at 520 nm. Draw a standard graph, and from this, the protein (sample) concentration is calculated.