Dual-Functionalized Porous Si/Hydrogel Hybrid for Label-Free Biosensing of Organophosphorus Compounds

A multifunctional porous Si (PSi) nanostructure is designed to combine a responsive PSi/hydrogel hybrid interfaced with a biorecognition element to selectively recognize small model molecules, organophosphorus compounds (OPCs), of high biological importance. A pH-responsive poly­(2-dimethylaminoethyl methacrylate) [poly­(DMAEMA)] hydrogel is synthesized and patterned in situ within an oxidized PSi Fabry–Pérot thin film. The resulting new hybrid displays a well-defined, rapid, and reversible optical response to pH changes. We employ this hybrid as an optical transducer element in a biosensing scheme by integrating it with organophosphorus hydrolase (OPH), capable of selective OPC hydrolysis. The enzyme is immobilized onto the pore walls of the oxidized PSi scaffold, resulting in an array of catalytic nanoscale chambers for the degradation of OPCs. Thus, the biosensor function relies on diffusion of the OPCs hydrolysis products from the catalytic chambers, through the interconnected pores network, to the hybrid region triggering its optical response. Exposure to the model target analyte results in a rapid and reproducible change in the optical reflectivity spectrum of the hybrid, allowing for label-free detection and quantification of OPCs in a simple and reliable manner

Dual-Functionalized Porous Si/Hydrogel Hybrid for Label-Free Biosensing of Organophosphorus Compounds

A multifunctional porous Si (PSi) nanostructure is designed to combine a responsive PSi/hydrogel hybrid interfaced with a biorecognition element to selectively recognize small model molecules, organophosphorus compounds (OPCs), of high biological importance. A pH-responsive poly­(2-dimethylaminoethyl methacrylate) [poly­(DMAEMA)] hydrogel is synthesized and patterned in situ within an oxidized PSi Fabry–Pérot thin film. The resulting new hybrid displays a well-defined, rapid, and reversible optical response to pH changes. We employ this hybrid as an optical transducer element in a biosensing scheme by integrating it with organophosphorus hydrolase (OPH), capable of selective OPC hydrolysis. The enzyme is immobilized onto the pore walls of the oxidized PSi scaffold, resulting in an array of catalytic nanoscale chambers for the degradation of OPCs. Thus, the biosensor function relies on diffusion of the OPCs hydrolysis products from the catalytic chambers, through the interconnected pores network, to the hybrid region triggering its optical response. Exposure to the model target analyte results in a rapid and reproducible change in the optical reflectivity spectrum of the hybrid, allowing for label-free detection and quantification of OPCs in a simple and reliable manner

Tethered Lipid Bilayers within Porous Si Nanostructures: A Platform for (Optical) Real-Time Monitoring of Membrane-Associated Processes

The importance of cell membranes in biological systems has prompted the development of artificial lipid bilayers, which can mimic the cellular membrane structure. Supported lipid bilayers (SLBs) have emerged as a promising avenue for studying basic membrane processes and for possible biotechnological applications. Conventional methods for SLB formation involve the spreading of lipid vesicles on hydrophilic solid supports. Herein, a facile approach for the construction of tethered SLB within an oxidized porous Si (pSiO₂) nanostructure, avoiding liposome preparation, is presented. We employ a two-step lipid self-assembly process, in which a first lipid layer is tethered to the pore walls resulting in a highly stable monolayer. A subsequent solvent exchange step induces the self-assembly of the unbound lipids into a robust SLB. Formation of pSiO₂–SLB is confirmed by fluorescence resonance energy transfer (FRET), and the properties of the confined SLB are characterized by environment-sensitive fluorophores. The unique optical properties of the pSiO₂ support are employed to monitor in real time the partitioning of a model amphiphilic molecule within the SLB via reflective interferometric Fourier transform spectroscopy (RIFTS) method. These self-reporting SLB platforms provide a highly generic approach for bottom-up construction of complex lipid architectures for performing biological assays at the micro- and nanoscale

Nanostructured Porous Si Optical Biosensors: Effect of Thermal Oxidation on Their Performance and Properties

The influence of thermal oxidation conditions on the performance of porous Si optical biosensors used for label-free and real-time monitoring of enzymatic activity is studied. 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

Label-Free Optical Biosensors Based on Aptamer-Functionalized Porous Silicon Scaffolds

A proof-of-concept for a label-free and reagentless optical biosensing platform based on nanostructured porous silicon (PSi) and aptamers is presented in this work. Aptamers are oligonucleotides (single-stranded DNA or RNA) that can bind their targets with high affinity and specificity, making them excellent recognition elements for biosensor design. Here we describe the fabrication and characterization of aptamer-conjugated PSi biosensors, where a previously characterized his-tag binding aptamer (6H7) is used as model system. Exposure of the aptamer-functionalized PSi to the target proteins as well as to complex fluids (i.e., bacteria lysates containing target proteins) results in robust and well-defined changes in the PSi optical interference spectrum, ascribed to specific aptamer-protein binding events occurring within the nanoscale pores, monitored in real time. The biosensors show exceptional stability and can be easily regenerated by a short rinsing step for multiple biosensing analyses. This proof-of-concept study demonstrates the possibility of designing highly stable and specific label-free optical PSi biosensors, employing aptamers as capture probes, holding immense potential for application in detection of a broad range of targets, in a simple yet reliable manner

Highly-Tunable Polymer/Carbon Nanotubes Systems: Preserving Dispersion Architecture in Solid Composites via Rapid Microfiltration

This research presents a new fabrication method for tailoring polymer/carbon nanotube (CNT) nanostructures with controlled architecture and composition. The CNTs are finely dispersed in polymeric latex, that is, polyacrylate, via ultrasonication, followed by a microfiltration process. The latter step allows preserving the homogeneous dispersion structure in the resulting solid nanocomposite. The combination of microfiltration and proper choice of the polymer latex, particle size, and composition allows the design of complex nanostructures with tunable properties, for example, porosity and mechanical properties. An important attribute of this methodology is the ability to tailor any desired composition of polymer–CNT systems, that is, nanotube content can practically vary anywhere between 0 to 100 wt %. Thus, for the first time, a given polymer/CNT system is studied over the entire CNTs composition, resembling two-phase polymer blends. The polyacrylate in these systems exhibits a structural transition from a continuous matrix (nanocomposite) to segregated domains dispersed within a porous CNTs network. An analogy of this structural transition to phase inversion phenomena in two-phase polymer blends is suggested. The resulting polyacrylate/CNT layers exhibit a percolation threshold as low as 0.04 wt %. Additionally, these nanomaterials show low total reflectance values throughout the visible, NIR and SWIR spectrum at a CNT content as low as 1 wt %, demonstrating their potential applicability for optical devices