Glucose Chemiluminescence Biosensor Based on Covalent Immobilization of Enzyme in Glutaraldehyde-Functionalized Glass Cell and Direct Coupling of Chitosan-Induced Au/Ag alloy Nanoparticles

Abstract

We report herein the development of a novel glucose chemiluminescence (CL) biosensor based on covalent immobilization of glucose oxidase (GOD) in glutaraldehyde-functionalized glass cell and direct coupling of chitosan-induced Au/Ag alloy nanoparticles on it, and how it may be useful for determination of glucose due to CL detection of enzymatically generated H₂O₂. In addition, the nanoalloy offers excellent catalytic activity toward hydrogen peroxide generation in enzymatic reaction between GOD and glucose and increases stability of covalent-linked enzyme. Chitosan molecules act as both the reducing and stabilizing agents for the preparation of nanoparticles (NPs) and also as a coupling agent between GOD and Au/Ag alloy NPs, which made possible the fabrication of a sensitive, accurate and stable biosensor for glucose. Under the optimum conditions, the biosensor can be used for the determination of glucose in the range of 1.2 × 10^–6 to 6.25 × 10^–3 M with a detection limit of 5.0 × 10^–7 M. The CL biosensor exhibited good storage stability, i.e., 90% of its initial response was retained after 2 months storage at pH 7.0. The present CL biosensor has been used to determine glucose in real serum and urine samples and validated against colorimetric spectrophotometry method.