In this study, we have modified glassy carbon electrodes (GCE) with amine groups (-NH2) to produce aminated GCE that can effectively detect current changes during urea hydrolysis. The silk fibroin (SF) scaffolds are employed to place enzyme near the surface of the aminated GCE. The as-fabricated electrode is employed for electrochemical urea sensing that could be monitored via cyclic voltammetry and amperometric techniques. Our developed biosensor has displayed rapid detection response (~1 min) and sensitivity (112.3 µA mM-1 cm-2) with linear correlation (0.3?8.4 mM) between the current and urea concentrations. Moreover, the analogous sensing responses obtained via SF scaffold discs (generated and functionalized) in the urease/SF/aminated GCE assuring the suitable platform for the urea sensing application devices.