In-device enzyme immobilization: wafer-level fabrication of an integrated glucose sensor

Wafer-level fabrication of integrated enzyme-based BioMEMS usually requires high temperature wafer-bonding techniques such as anodic bonding. Enzymes denature at comparatively low temperatures. Thus, enzymes need to be immobilized after wafer bonding. A convenient in-device immobilization method is presented allowing wafer-level patterning of enzymes inside micro-scale flow channels after wafer bonding. Enzymes are entrapped in a poly(vinyl alcohol)-styrylpyridinium (PVA-SbQ) membrane crosslinked by UV exposure through a transparent top wafer. The reaction kinetics of immobilized glucose oxidase is investigated in more detail. A low apparent Michaelis constant of 3.0 mM is determined indicating a rapid diffusion of glucose into the PVA-SbQ membrane as well as an oxygen-limited maximum catalytic rate. The entrapped glucose oxidase preserves its native properties since it is not chemically modified. Furthermore, the active PVA-SbQ membrane can be dehydrated in a vacuum and later rehydrated in buffer solution without significant loss of enzyme activity. An integrated enzyme-based glucose sensor fabricated on a wafer-level using in-device immobilization is described to demonstrate the potential of this novel technique. The sensor is part of a disposable microneedle-based continuous glucose monitor. The stability of glucose oxidase entrapped in PVA-SbQ is sufficient to continuously operate the sensor at 25 °C for 24 h.