Apoptosis is a crucial event for the efficacy of anticancer drug. Caspase-3 plays a critical role in cell signaling pathways of apoptosis. The calixarene derivatives display high affinity for various kinds of biomolecules by host-guest recognition own to their unique cavity structure. The calixarene functionalized reduction of graphene oxide exhibits stronger molecular recognition ability and a higher electrochemical response to biomolecules than unmodified graphene oxide. In this work, caspase-3 recognizes and cleaves N-terminal blocked peptide containing tetra-peptide substrate Asp-Glu-Val-Asp. p-sulfonatocalix[6]arenes sodium modified graphene oxide (pSC6-rGO) recognizes exposed N-terminal amine group assembled on the electrode. Due to the large surface area to volume ratio of rGO, numerous electrochemical active methylene blues (MB) can be absorbed through host-guest recognition. As a result, sensitive caspase-3 detection was achieved with a low detection limit of 0.0167 pg/mL by this electrochemical signal amplification strategy. This approach was also applied to measure apoptosis in the practical cell samples and shows good performance.