Neuropathy Target Esterase (NTE) is a membrane protein found in human neurons, and other cells, including lymphocytes. Binding of certain organophosphorus (OP) compounds to NTE is believed to cause OP-induced delayed neuropathy (OPIDN), a type of paralysis for which there is no effective treatment. Mutations in NTE have also been linked with serious neurological diseases, such as Lou Gehrig's disease. In this paper, for the first time, a nanostructured biosensor containing a domain of NTE known as NEST has been developed. The biosensor has a response time on the order of seconds and gives dose-dependent decrease in sensor output in response to known NEST (or NTE) inhibitors. Potential applications of the biosensor include studying the fundamental reaction kinetics of NTE mutants, screening OP compounds for NTE inhibition, and investigating the effect of NTE mutations on NTE esterase activity. While the development of NEST biosensor was the primary purpose of this study, we found that the approach that we have developed for NEST can also easily be extended to immobilize and measure the activity of other medically relevant esterases such as Acetylcholinesterase and Butyrylcholinesterase. Electrochemical impedance spectroscopy (EIS), potential step voltammetry and ellipsometry were the primary characterization technique used in this study.