(Invited) Application of High Hydrostatic Pressure and Nanofilms to the Fabrication of Enzyme Biosensors

Abstract

Fabrication of enzyme biosensors has evolved since they were first conceived in the late 1950s. Increased sensitivity and reproducibility have been perhaps the two main driving forces for the development of such sensors. Increased sensitivity enables miniaturization and the development of implantable sensors for in-vivo applications. Because most oxidoreductases used in the fabrication of enzyme biosensors quickly lose their activity over time, most enzyme biosensors either are one-use disposable strips or require periodic change of membrane and frequent calibration, which makes them impractical for industrial applications. With the nanotechnology boom, several research reports have focused on increased sensitivity by creating porous nanostructures. There are also several reports on the effects of immobilization, cross-linking, and additions of solutes to stabilize enzymes and extend the operational life of enzyme biosensors. Here we summarize our research for the last 10 years on the effects of platinization conditions, including the use of high hydrostatic pressure (HHP) up to 500 MPa on the fabrication of microporous platinum black structures that maximize electrode sensitivity. In our early research we reported a 60-fold increase in sensitivity of glucose biosensors. We also report the effect of enzyme entrapment in poly-o-phenylenediamine (PoPD) nanofilms and their impact on electrode sensitivity and stability on glucose oxidase, alcohol oxidase, and galactose oxidase. High pressure and immobilization in PoPD resulted in two orders of magnitude increase in glucose oxidase biosensors. In contrast, while HHP increased the stability of AOx by a factor of ~15, immobilization only increased its stability by a factor of ~5 stability and combined HHP and immobilization only increased the stability by a factor of 6.4 relative to the enzyme in solution.