Amperometric biosensor with enzyme amplification fabricated using self-assembled monolayers of alkanethiols: the influence of the spatial distribution of the enzymes

The influence of the spatial distribution of the two enzymes in an amplified enzyme electrode is investigated by the step-wise fabrication of the enzyme interface. The two enzymes, glucose oxidase (GOD) and glucose dehydrogenase (GDH), are co-immobilised onto the surface of a gold electrode modified with the alkanethiol, 3-mercaptopropionic acid. Three geometries were fabricated: (1) a monolayer enzyme electrode where both enzymes were covalently attached to the alkanethiol layer, (2) a bilayer electrode where the inner layer was GOD and the outer layer was GDH, and (3) a bilayer sensor where the inner layer was GDH and the outer layer was GOD. The response of the three geometries was shown to vary with regard to linear range, sensitivity and hence gain. We believe this is the first demonstration of the spatial relationship between enzymes in a multi-enzyme system influencing the response of the resultant enzyme electrode.