Determination of conjugated protein on nanoparticles by an adaptation of the Coomassie blue dye method

A new accurate spectrophotometric method for protein determination on nanoparticles is described. The method is based on the Coomassie blue dye that binds to the basic and aromatic amino acid residues of proteins, especially arginine and lysine. A known amount of reagent dye was mixed with a variety of protein-loaded nanoparticles. Thereafter the unconjugated reagent was mixed with excess protein (bovine serum albumin) and titrated. In this method, the reacted dye on the protein coating of nanoparticle is directly determined, in opposite to the conventional method, in which the conjugated protein is determined as the difference between the non-conjugated protein found in the supernatant after centrifugation, and the total amount of protein originally used. This method is able to measure amounts of coated protein lower than 1 ppm. Simple and accurate method especially adapted for protein-coated nanoparticles. The amino acid residues of protein in the nanoparticle surface react with Coomassie brilliant blue dye. The unreacted dye is titrated with an excess of a standard protein. (C) 2019 The Author(s). Published by Elsevier B.V.