An amperometric thin-film glucose biosensor based on a plasma-polymerized film using hexamethyldisiloxane as the monomer is presented. The plasma-polymerized film, achieved in plasma in the vapor phase, offers a new alternative for use in the design of the electrode-enzyme interface of biosensors. The film shows promise of high sensor performance; namely, rapid sensor response, low noise, a wide dynamic range, reproducibility, and reduction in the effects of interfering materials such as ascorbic acid. In this study, we examined the usefulness of the hexamethyldisiloxane plasma-polymerized film and investigated how the thickness of the plasma-polymerized film on a platinum electrode affected sensor characteristics: the selectivity for hydrogen peroxide versus interfering agents, the sensor response due to enzymatic reaction, and oxygen depletion.