A novel signal amplification method for surface plasmon resonance (SPR) was developed to detect Caspase-3 based on cucurbit[7]urils (CB[7]) and bimetallic nanonetwork. Three-dimensional (3D) bimetallic nanonetwork was formed through CB[7]-integrated self-assembly of gold nanoparticles (AuNPs) and silver nanoparticles (AgNPs) (AuNPs-CB[7]-AgNPs) by one-pot synthesis method. Caspase-3 was utilized as a model protein and peptide EEAAADEVDFKKAAAC was used as the substrate. In the presence of Caspase-3, peptide was hydrolyzed and exposed phenylalanine at the N-terminus, which could bind to CB[7] through host-guest interaction. The multivalent receptors CB[7] on bimetallic nanonetwork provide a simple strategy to increase the binding affinities between 3D bimetallic nanonetwork and peptide. Thus, AuNPs-CB[7]-AgNPs could attach to the gold chip with ease. Remarkably, due to the strong optical properties and high refractive index of 3D bimetallic nanonetwork, it has a higher SPR signal enhancement than that of only AuNPs or AgNPs. Therefore, a highly sensitive Caspase-3 sensing strategy could be achieved with a low detection limit of 5.39 pg mL−1, providing a new and facile SPR signal amplification method for detection of cell apoptosis.