Peroxidase-Like Activity of Gold Nanoparticles and Their Gold Staining Enhanced ELISA Application

We found that the peroxidase-like activity of gold nanoparticles (GNPs) followed the Michaelis-Menten kinetic model and was dependent on environmental pH and temperature, which was very similar to natural Horseradish Peroxidase (HRP). However, unlike HRP, which needs a lower H2O2 concentration with a very narrow range to reach a maximum reaction rate and avoid enzyme poisoning, GNPs have very high activity, even at an H2O2 concentration two orders of magnitude higher than HRP. It was demonstrated that H2O2 treatment could enhance the peroxidase-like activity of GNPs, resulting thus in the activity increase in a circular catalytic reaction by the reduplicative use of GNPs. It was also found that the peroxidase-like activity of GNPs responded sensitively to nanoparticle size and surface modifications. When used in an immunoassay, GNPs were generally conjugated with antibody and blocked with hydrophilic macromolecules to construct a nanoprobe. This strongly reduced the peroxidase-like activity and detection sensitivity of GNPs, therefore, restricting their use as peroxidase mimetics. We presented a novel strategy that combined the nanoprobes with gold staining to expose fresh catalytic gold surfaces and obtained a great increase in detection sensitivity.