Controlled deposition of glucose oxidase on platinum electrode based on an avidin/biotin system for the regulation of output current of glucose sensors.

A facile method for the regulation of enzyme loading on an electrode surface has been studied using avidin and biotinylated glucose oxidase (GOx). It was demonstrated that an alternate and repeated deposition of avidin and biotinylated GOx gives a protein thin film probably composed of avidin monolayers and biotinylated GOx monolayers which are connected with each other through strong affinity between avidin and biotin moieties of the enzyme (binding constant, 10(15) M-1). Amperometric response of the glucose sensors constructed by this method was controlled stepwise and rather precisely by regulating the number of GOx layers deposited (or the loading of GOx). For example, the output current of the sensors to 1 mM glucose was enhanced to ca. 1300 and 2800 nA after deposition of 10 and 20 layers of GOx, respectively, as compared with 110 nA for the monolayer GOx sensor. The enhanced response contributed to the extension of the dynamic range of the sensors, especially at a lower glucose concentration. The response time of the sensors was satisfactorily fast (ca. 20 s), irrespective of the number of GOx layers.