Layer-by-layer immobilization of horseradish peroxidase on a gold electrode modified with colloidal gold nanoparticles

An amperometric biosensor, based on layer-by-layer self-assembly of colloidal gold nanoparticles, cysteine and horseradish peroxidase on Nafion modified electrode surface by electrostatic adsorption, has been used for the determination of hydrogen peroxide. The electrochemical behavior of the multilayer film was studied by cyclic voltammetry, linear sweep voltammetry and chronoamperometry. The step layer-by-layer adsorption interface morphology was further characterized by means of electrochemical impedance spectroscopy and cyclic voltammetry. The performance and factors influencing the resulted biosensor were studied in detail. The sensor displayed an excellent electrocatalytic response to the reduction of H2O2 without the aid of an electron transfer mediator. Linear response to H2O2 was obtained for the concentration range from 1.6 mu M to 2.4 mu M under optimized conditions. The detection limit of the biosensor was 0.5 mu M (S/N = 3), and the sensor achieved 95% of the steady-state current within 10 s. The sensor exhibited high sensitivity, selectivity and stability.