Immobilized biomolecules are widely applied in medical, food and environmental fields. In this paper, glucose oxidase (GOD) was covalently immobilized on the surface of thiol-modified silver nanoparticles (AgNPs) to fabricate GOD-AgNP bioconjugate complexes. GOD-AgNP complex were more stable at high temperature and in high pH environments than the free GOD in solution. Additionally, the fabricated Carbon rod/GOD-AgNPs/Nafion/Chitosan electrode exhibited a rapid response and a linear calibration range from 0.5 mM to 6 mM for the detection of glucose. The Nafion/Chitosan coating could eliminate common interferents, such as uric acid and acetaminophen. As a conclusion, the experimental results indicated that the resulting modified electrode possesses an excellent response performance on glucose.