Abstract Microelectrode array (MEA) has recently attracted significant research interest in biomedical applications due to its high sensitivity, mass production and fast response. However, the previous MEA with precious metal-based nanomaterials for electro-catalysis may induce toxicity to cells. In this work, we present a metal-free graphene/graphene oxide (rGO/GO) hybrid MEA for precise and in-situ H2O2 secretion detection. The configuration and size of rGO/GO hybrid MEA can be well designed and controlled by combining the convenient lithography and electrochemistry method, where the GO region with abundant oxygen functionalized groups has good biocompatibility, so as to play as the adhesion and growth agent of pheochromocytoma cells (PC12 cells). Whereas, the rGO region with higher electro-catalytic activity can favor high sensitive in-situ H2O2 secretion detection that is released from the stimulated PC 12 cells at GO region, but with the cell filopodia extended to the rGO region. Besides, the rGO/GO hybrid MEA presented herein is compatible with the micro-fabrication technique and can be integrated into the lab-on-chip devices, which may shed useful insights for the design and fabrication of high performance biosensors.