Advances in Glucose Oxidase-Based Electrochemical Biosensors: Generational Development, Immobilization Strategies, and Nanostructure Applications

Monitoring glucose levels is vital for the management of diabetes and other metabolic disorders, spurring extensive research into the development of reliable, sensitive and biocompatible glucose sensors. Glucose oxidase (GOx)-based electrochemical biosensors have evolved over time through three different generations, with each generation aiming to overcome the limitations of the previous one. First-generation sensors depend on oxygen as the electron carrier, while second-generation systems utilize artificial mediators that provide more stable and sensitive measurements at lower potentials. Third-generation sensors aim to enable direct electron transfer (DET) between the enzyme and the electrode, enabling the development of marker-free, biocompatible platforms suitable for continuous glucose monitoring. Enzyme immobilization methods such as physical adsorption, covalent binding, entrapment in polymer matrices, crosslinking and microencapsulation play a critical role in improving sensor performance, stability and reproducibility. Moreover, the integration of nanostructures such as graphene, carbon nanotubes, gold nanoparticles and hybrid composites on the electrode surface facilitates efficient electron transfer and improves sensor response by increasing surface area, conductivity and biocompatibility. This review provides a comprehensive overview of the basic principles, developmental processes and challenges of GOx-based glucose biosensors, with a particular focus on recent advances in enzyme immobilization strategies and nanostructured electrode materials. The integration of these approaches is anticipated to contribute significantly to the development of wearable, noninvasive, and continuous glucose monitoring systems, which are of great importance for personalized healthcare and diabetes management.