Diabetes is a chronic disease, which can give serious damages to the human organs that affect life expectancy. Such a life-threatening diabetes is diagnosed by systematic monitoring of blood glucose levels; thus, accurate detection of glucose within a specified target range becomes more important for glucose sensor to provide detailed information relating with diabetes, which can result in reliable decision for diabetes treatment. Thanks to many efforts developing the reliable glucose sensors, electrochemical performance of the sensor including sensitivity and selectivity in response to glucose is gradually improved. Herein, we developed spinel type NiCo2O4 (NCO) nanostructure with excellent sensitivity, selectivity and chemical robustness for glucose detection. NCO was prepared by conversion of NiCo-layered double hydroxide (NCH) at comparatively low temperature, which is synthesized by simple and facile chemical bath method and following post-heat treatment of the NCH. Electrochemical sensitivity, selectivity and detection time of the NCO in response to glucose was investigated, compared to those of the NCH. Also, long-term stability of the NCO on repetitive glucose detection was evaluated. Based on the systematic analysis on materials properties and electrochemical performance of the NCO, possible mechanism of the glucose oxidation, which significantly improves electrochemical performance of the NCO, is discussed.