Modification of electrode surface for covalent immobilization of laccase

Various methods for covalent immobilization of laccase on electrode surface were tested. In case of glassy carbon (GC) electrode, the surface was modified by electrochemical oxidation of 1,5-pentanediol or by direct electrochemical oxidation of the electrode itself to introduce carboxylic functional group. The peptide coupling between laccase and the functional groups of the modified electrode was done by the use of 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride and N-hydroxysuccinimide (EDC/NHS). The direct modification method introduces denser carboxylic functional group. In the case of platinum (Pt) electrode, the surface was modified by direct electrochemical oxidation to introduce hydroxy functional group, which was followed by coupling the enzyme with cyanuric chloride (CC). Another method tested was the modification of the surface by silanization with 3-aminopropyltriethoxysilane (APTES), which was followed by coupling the enzyme with glutaraldehyde (GA). Among the above four types of modification, the silanization method is the most effective with respect to long-term stability and fast response for biosensor uses.