Cobalt oxide (Co3O4)-based materials have been extensively investigated as efficient electrocatalysts for non-enzymatic glucose biosensing. The proper tuning the structure of Co3O4-based materials could improve sensing performances for glucose detection. Herein, we have developed a combined self assembly and calcination method to prepare nanoparticles-assembled Co3O4 nanosheets, which exhibit good sensing performances for glucose biosensing. The Co3O4 nanosheets were prepared by the following three steps: (i) the Co2+ modified graphene oxide (GO) was prepared by the self-assembly of Co2+ and GO in aqueous solution firstly, and then Co2+ modified GO film was obtained by vacuum filtration method; (ii) after calcination of Co2+ modified GO film in N-2 atmosphere, Co3O4 nanoparticles modified reduced GO (RGO) was obtained; (iii) finally, nanoparticles-assembled Co3O4 nanosheets were obtained by calcination of Co3O4 modified RGO in air to remove RGO template. Most importantly, Co3O4 nanosheets exhibit good electrocatalytic activity for oxidation of glucose, leading to high-performance glucose sensor. The detection limit and linear range of Co3O4 nanosheets-based glucose sensor in the present work are 0.15 mu M, and 1-50 mu M, respectively. Furthermore, such sensor also shows excellent selectivity for glucose detection, compared to the commonly interfering species including dopamine, and uric acid. (C) 2016 Elsevier Inc. All rights reserved.