Three-dimensional (3D) bismuth selenide (Bi2Se3) nanostructures were synthesized by microwave synthesis using water as a solvent and hydrazine hydrate as a reducing agent and exfoliated into few layers of Bi2Se3. Bi2Se3- Few Layers (Bi2Se3- FL) exhibited localized surface plasmon resonance and enhanced electrocatalytic behavior. The scanning electron microscope (SEM) and transmission electron microscopy (TEM) characterization indicated the layered structure of Bi2Se3. The electrocatalytic properties of the Bi2Se3-FLmodified GC electrode towards non-enzymatic glucose oxidation were evaluated by cyclic voltammetry (CV) and chronoamperometry. The designed non-enzymatic glucose sensor showed a low detection limit of 6.1 μM, a linear range from 10 μM to 100 μM of glucose concentration and a current sensitivity of 0.112 μAμM−1.The electrochemical sensor constructed using Bi2Se3-FL attained steady-state level within 3 s upon adding glucose and remained stable even after 19 days with only 17% loss in current signal. The obtained electrodes can be applied for determining glucose in urine samples. The results obtained here are of great significance to use nanostructured Bi2Se3-FL electrode as a potential candidate for non-enzymatic glucose detection.