(41e) Electrochemical Detection of Enzyme Kinetics Using Nanofluidic Devices

Nanoscale devices are interesting because they approach the size scale of individual molecules and offer the opportunity for investigating biological systems from new physical perspectives. We have demonstrated solid-state devices consisting of a 50 nm high solution-filled cavity bounded by two parallel electrodes in which electrochemically active molecules transfer charge between suitably biased electrodes. Electrochemical reactions typically only involve one or a few electrons per molecule rendering direct detection of a single molecule virtually impossible. By utilizing electrochemically active molecules in our setup, which can repeatedly undergo reversible reduction and oxidation, each molecule can transfer, on average, thousands of electrons between the electrodes while residing in the cavity.

This talk will focus on employing these devices for determining enzyme kinetics. The reaction kinetics can be monitored directly by the device if the enzyme is capable of converting an electrochemically inactive substrate into an active product. Since the nanochannel is open to the bulk, the composition of the solution inside the device is a representative sample of the bulk solution. Therefore, as the concentration of product in the bulk solution increases, the average number of product molecules in the nanochannel also goes up leading to a larger signal. Our model enzyme is tyrosinase, which converts electrochemically inactive monophenols into active diphenols and quinones. We have observed the conversion of different monophenol substrates by tyrosinase and compared the results against standard UV-Vis spectroscopy techniques. This fluidic approach reduces the sample volume and enzyme concentrations needed for these types of experiments. More importantly, it serves as a foundation for more complex biophysics experiments and integrated fluidic systems.