The glucose electrooxidation reaction at NiOx nanoparticles loaded on a nitrogen-doped carbon nanofiber (N-CNF)-modified glassy carbon electrode (NiOx@N-CNF/GCE) in an alkaline medium was studied. The N-CNF was produced by an electrospinning technique to acquire a large surface area. The produced electrode was characterized by X-ray diffraction (XRD) and scanning electron microscopy (SEM) coupled with energy-dispersive X-ray (EDX) analysis. The electrocatalytic properties of this electrode towards glucose oxidation were tested by cyclic voltammetry and linear sweep voltammetry (LSV). T he NiOx@N-CNF/GCE catalytic activity towards glucose oxidation in an alkaline medium was found to be excellent in terms of measured glucose concentration and electrode stability. It showed high reproducibility and stability towards glucose oxidation with activity r etention e ven a fter 1 00 c ycles o f c ontinuous p otential s canning. I n a ddition, N iOx@ N-CNF/GCE showed a good linear behavior of glucose concentration detection in the concentration range between 0.0 and 10.0 mM, which highlights its ability to be used as a nonenzymatic glucose biosensor.