Imprinted monolayers have several advantages over bulk imprinted polymers such as excellent mass transfer of molecules into and out of imprinted sites and transduction of binding signals detected in real time. Protein-imprinted self-assembled monolayers (SAMs) were created with multiple binding sites and biocompatible imprinted cavities from functional thiols and novel disulfide compounds containing an oligoethylene glycol (OEG) terminal moiety and two amide groups incorporated in the chain (DHAP) in a biologically benign solution. DHAP played an important role in the formation of multiple binding sites and biocompatible cavities in addition to resisting nonspecific protein binding. The created protein-imprinted SAMs exhibited the excellent ability of specific binding of target proteins determined by multiple binding sites and imprinted cavities. The strategy generates tailor-made monolayer surfaces with specific protein binding and opens the possibility of controlled assembly of intellectual biomaterials and preparation of biosensors.