In the present work, a glucose sensor was developed based on the glucose oxidase immobilized on Pd (Pd-GOx) and nafion modified electrode. The modified electrode was characterized by atomic force microscopy and field emission scanning electron microcopy. The electrochemical analysis for electrochemical activity was accessed by cyclic voltammetry. The modified electrode displayed an excellent electrooxidation behavior to glucose and was detected by cyclic voltammetry and amperometry. In optimized conditions, the fabricated Nafion/Pd-GOx modified electrode exhibited a sharp amperometric response to Human blood and glucose. The modified electrode also holds its high selectivity in the presence of bilirubin, creatinine, and uric acid, indicating it can be an ideal electrode material for detection of glucose in diabetes.