Adsorption of glucose oxidase (GOD) onto plasma-polymerized thin films (PPF) with nanoscale thickness was characterized by atomic force microscopy (AFM), quartz crystal microbalance (QCM), and electrochemical measurements. The PPF surface is very flat (less than 1-nm roughness), and its properties (charge and wettability) can be easily changed while retaining the backbone structure. We focused on three types of surfaces: (1) the pristine surface of hexamethyldisiloxane (HMDS) PPF (hydrophobic and neutral surface), (2) an HMDS PPF surface with nitrogen-plasma treatment (hydrophilic and positive-charged surface), and (3) an HMDS PPF surface treated with oxygen plasma (hydrophilic and negative-charged surface). The AFM image showed that the GOD molecules were densely adsorbed onto surface 2 and that individual GOD molecules could be observed. The longer axis of GOD ellipsoid molecules were aligned parallel to the surface, called the "lying position", because of electrostatic association. On surface 1, clusters of GOD molecules did not completely cover the original PPF surface (surface coverage was ca. 60%). The 10-nm-size step height between the GOD clusters and the PPF surface suggests that the longer axes of individual GOD molecules were aligned perpendicular to the surface, called the "standing position". On surface 3, only a few of the GOD molecules were adsorbed because of electrostatic repulsion. These results indicate that the plasma polymerization process can facilitate enhancement or reduction of protein adsorption. The AFM images show a corresponding tendency with the QCM profiles. The QCM data indicate that the adsorption behavior obeys the Langmuir isotherm equation. The amperometric biosensor characteristics of the GOD-adsorbed PPF on a platinum electrode showed an increment in the current because of enzymatic reaction with glucose addition, indicating that enzyme activity was mostly retained in spite of irreversible adsorption.

We describe a method for creating a mediator-containing interface between an enzyme and an electrode, achieving simpler and more reliable immobilization of the enzyme with the enhanced detection sensitivity. A nanothin polymer film containing a redox mediator, made of dimethylaminomethylferrocene, was plasma-deposited directly onto a glucose oxidase-physisorbed electrode, with which a relevant bioelectrochemical signal was observed without prior or further chemical modification of the enzyme molecules. The results of the surface characterizations before and after the enzyme immobilization showed that this method gave control over the spatial orientation of single enzyme molecules in favor of efficient and reproducible signal generation. Considering that the film deposition was performed using microfabrication-compatible organic plasma, our new method has a great potential of enabling high-throughput production of bioelectronic devices without chemical modification steps.

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We propose an electron transfer-mediated amperometric enzyme biosensor based on plasma-polymerized thin film of dimethylaminomethylferrocene (DMAMF) on a sputtered gold electrode. The DMAMF plasma-polymerized film is deposited directly onto the surface of the electrode under dry conditions. The resulting thin film not only has redox sites but also is extremely thin (∼20 nm), adheres well onto the substrate (electrode), has a flat surface and a highly-crosslinked network structure, and is hydrophilic in nature. Glucose oxidase is densely immobilized onto the surface of DMAMF plasma-polymerized film on the gold electrode. From the electrochemical measurement, the biosensor can cover the wide range of glucose concentration (1.3 -81 mM) at +350 mV of applied potential. The current response of the glucose biosensor was decreased by less than 5% in an aerobic solution as compared to that in an anaerobic solution. These show that the DMAMF plasma-polymerized films play a role as the electron transfer mediators between the reaction center of enzyme and the electrode.

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