Giorgi Shtenberg1

1, ARO Volcani Center, Bet Dagan, , Israel

Nanostructured porous Si (PSi) has emerged as an attractive and versatile material for optical biosensing applications due to its large internal surface area and tunable optical properties. Numerous biosensing schemes have been reported, demonstrating the advantages of these nanosystems over conventional bio-analytical techniques in terms of improved detection sensitivity, label-free, and real-time rapid analysis. However, a key challenge in designing PSi-based biosensors arises from the relative chemical instability of the Si scaffold in biologically-relevant environments. Specifically, PSi oxidation and dissolution in aqueous environments lead to significant changes in its optical and electrical properties, e.g., luminescence, refractive index and absorption coefficient, and may ultimately result in the structural collapse of the matrix. Several chemical routes are used to enhance PSi stability, including thermal oxidation, hydrosilylation, electrochemical alkylation, and thermal hydrocarbonization. Thermal oxidation is frequently utilized for PSi biosensors passivation, owing to the wide repertoire of chemical modifications available for Si oxide surfaces. The present work explores the effect of thermal oxidation conditions on the stability and sensitivity of PSi-based optical biosensor for label-free and real-time monitoring of enzymatic activity. We compare three oxidation temperatures (400, 600, and 800 °C) and their effect on the enzyme immobilization efficiency and the intrinsic stability of the resulting oxidized porous Si (PSiO2), Fabry–Pérot thin films. Importantly, we show that the thermal oxidation profoundly affects the biosensing performance in terms of greater optical sensitivity, by monitoring the catalytic activity of horseradish peroxidase and trypsin-immobilized PSiO2. Despite the significant decrease in porous volume and specific surface area (confirmed by nitrogen gas adsorption–desorption studies) with elevating the oxidation temperature, higher content and surface coverage of the immobilized enzymes is attained. This in turn leads to greater optical stability and sensitivity of PSiO2 nanostructures. Specifically, films produced at 800 °C exhibit stable optical readout in aqueous buffers combined with superior biosensing performance. Thus, by proper control of the oxide layer formation, we can eliminate the aging effect, thus achieving efficient immobilization of different biomolecules, optical signal stability, and sensitivity.