S. Sayin, Y. Zhou, K.D. Benkstein, K.L. Steffens, S. Xin, S. Semancik, M. Zaghloul
The George Washington University,
Keywords: biosensors, plasmonic sensors, nanosensors, LSPR sensors, biomedical diagnostics
Surface plasmon resonance (SPR) is an optical technique used to measure refractive index changes in very thin layers adsorbed on certain metals. This phenomenon arises due to the interaction between incident light energy and delocalized electrons in the metal films, which results in a decrease in reflected light intensity at a sharp angle. Localized surface plasmon resonance (LSPR)-based nanosensors provide the possibility for real-time and label-free monitoring of immobilized probe/target binding phenomena. We are currently developing LSPR-based nanohole arrays (NHAs) as a sensing platform that utilizes optical signals derived from local "hot spots" where changes in the refractive index occur when molecules interact. The NHA-based sensors are advantageous due to their miniaturized features, high sensitivity, cost-effectiveness, and potential for point-of-care use. Our goal is to monitor low-level biomolecular target species relevant to biomedical diagnostics, such as the SARS-CoV-2 virus. To adapt these sensors for SARS-CoV-2 virus detection, we plan to use surface-immobilized nanobodies to capture the target molecule from solution-phase samples. Using a finite-difference time-domain simulator, we determined that 75 nm Au thickness is optimal for NHA structures. The structure of NHA was studied using scanning electron microscopy (SEM) and an observed image is shown in Figure 1(b). Our sensing system consists of a portable spectrometer, optical fibers, probe station, microscope, light source, and a newly designed sensor chamber. Previous work demonstrated the utility of the NHA sensor for gas-phase detection of small molecules, while in this work we demonstrate the design, fabrication, and testing of a PDMS microfluidic channel suitable for liquid-phase sensor operations. To create a PDMS well, a 3D printed mold was utilized. The mold design featured a stream channel for fluid passage, an inlet and outlet for fluid introduction and discharge, and an opening for sensor placement. The PDMS was produced with the mold, and then a glass slide was affixed to the bottom. After placing the four-sector sensor chip in the sensor opening, the top was sealed with a glass cover slip permitting sample handling through the inlet and outlet. This sensor chamber enabled reflectance mode measurements from each of the four sectors in the plasmonic platforms. To demonstrate the functionality of the sensor in the new sensor cell, a series of tests were conducted. These ranged from simple air/water tests to water:ethanol solutions with targeted refractive indices. In all cases, the LSPR peak position was monitored as a function of solution refractive index with the goal of demonstrating the sensor effectiveness and reproducibility in the new sensor cell and a liquid-phase environment. Functional surface chemistries involving PEGylation, BSA model protein and SARS-CoV-2 nanobody immobilization on the Au surfaces are also being developed to introduce target-specific capture probes for biomedical applications, where binding of a target biomolecule yields a local refractive index change and measurable signal. The surface functionalization steps on clean Au surfaces have been characterized by X-ray photoelectron spectroscopy (XPS). BSA attachment was used as a model protein attachment prior to nanobody-based studies.