Enhanced Glucose Sensing through Optimization of Glucose Oxidase and Osmium-Based Redox Polymer on Gold Electrodes

Glucose oxidase (GOx)-based electrodes offer promising applications in glucose sensing and as potential power sources for implantable devices, yet their performance remains critically dependent on efficient electron transfer and enzyme immobilization strategies. This study systematically investigated the co-immobilization of GOx and a redox-active osmium polymer, poly (N-vinylimidazole)-[Os(4,4′-dimethyl-2,2′-bipyridine)₂Cl])^+/2+ (PVI-Os-dme), using poly(ethylene glycol) diglycidyl ether (PEGDGE) as a crosslinker to enhance both the catalytic and electron-transfer properties of the electrode. By varying the enzyme-to-mediator ratio and applying a layer-by-layer assembly approach, we demonstrated that both loading quantity and composition critically influenced current generation, charge transfer resistance, and overall electrode efficiency. While current output increased with additional layers, the catalytic activity per unit mass of enzyme or mediator decreased, indicating a trade-off at high loadings. The optimized electrode, composed of six composite layers (2 μg GOx, 3.6 μg PVI-Os-dme, 2.2 μg PEGDGE per layer), achieved the highest peak current of 23.7 ± 1.7 μA at 0.3 V and retained over 85% of initial current after 3 cycles and 57% after 5 cycles, demonstrating favorable reusability. Kinetic analysis revealed an apparent Michaelis–Menten constant (K_m^app) of 9.0 mM and a maximum current (I_max) of 29.2 μA, confirming the electrode’s high affinity and catalytic efficiency toward glucose. These results highlight the importance of optimizing GOx/PVI-Os-dme loadings, ratio, and the number of layers for enhancing the electrode performance.