Membraneless immobilization of xanthine oxidase on organic conducting salt/silicone oil electrodes

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Abstract
Xanthine oxidase (XOD, E.C. 1.2.3.2) has been immobilized on amperometric electrodes made from the organic conducting salt tetrathiafulvalene‐p‐tetracyanoquinodimethane (TTF‐TCNQ) and silicone oil. Xanthine oxidase was immobilized by various techniques; immobilization by means of hydrophobic interaction was found to yield optimum analytical performance. Except for buffer, no coreactants are required for sensor operation. The response of the XOD electrode to its substrate hypoxanthine can be described by the enzyme‐kinetic Michaelis‐Menten formalism with excellent correlation (in most cases, Γ2 > 0.995). Steady‐state currents are usually measured within 10 seconds. Apparent KM (typically 3 to 10 μM) and saturation current values (0.2 to 3.7 μA, measured with new electrodes) strongly depend on the preparation procedure chosen as did the useful lifetime of the XOD sensors which was up to several weeks at room temperature. The described immobilization methodologies circumvent diffusion problems associated with well‐established biosensor preparation techniques (e.g., physical entrapment in gel or between dialysis membranes) and thus yield biosensors of superior analytical performance.