Colorimetric protease assay by using gold nanoparticles and oligopeptides

A colorimetric protease assay based on the cleavage of gold nanoparticles (AuNPs) aggregates cross-linked with oligopeptides is developed. In this assay, when oligopeptide YHPQMNPYTKAGGGC is added to a solution containing citrate-stabilized AuNPs, cysteine and lysine on the oligopeptide causes rapid aggregation of AuNPs, resulting in color change from red to blue. However, in the presence of a model protease trypsin, the oligopeptide is cleaved by trypsin into two fragments. As a result, AuNPs do not aggregate and no color change can be observed. By using this principle, trypsin can be detected with the naked eye at a concentration as low as 0.5 nM. This assay can also be applied to detect trypsin inhibitors such as benzamidine hydrochloride.