Biology subjects

Shih, W.-C.

Publications and source records attributed to Shih, W.-C..

3 recordsLinked to original sources

bioRxiv2026

Isolation and Purification-Free Digital Single-Small Extracellular Vesicle Biosensing with Scalable Plasmonic Arrays

Recent advances in plasmonic biosensing and imaging have enabled label-free analysis of single biological nanoparticles. We previously developed PlAsmonic NanOapeRture lAbel-free iMAging (PANORAMA) for isolation and purification-free, digital counting and precise localization of small extracellular vesicles (sEVs), with complementary fluorescence interrogation of surface and intravesicular biomarkers for quantitative molecular profiling. The fact that no isolation and purification or isolation is needed represents a crucial advantage because various specificity, efficiency, and time-consumption issues hinder quantitatively reproducible extraction of sEVs from biological fluids. PANORAMA achieves ultrahigh refractive-index sensitivity through arrayed gold nanodisks on invisible substrates (AGNIS) fabricated by nanosphere lithography (NSL). However, despite its simplicity and low cost, NSL is frequently constrained by poor large-area uniformity, which hinders scalable fabrication. Here, we introduce nanosphere settling lithography (NSSL) as an alternative to the gold-standard Langmuir-Blodgett trough (LBT) process, enabling highly uniform, large-area monolayers with reduced process stringency. AGNIS fabricated via NSSL exhibits high refractive-index sensitivity with low spatial variability across 60 mm x 24 mm substrates, sufficient for 60-well in standard 384-well plate format. The platform demonstrates exquisite sensitivity through PANORAMA digital counting and sizing of 25, 50, and 100 nm polystyrene beads, as well as single-vesicle characterization of sEVs derived from H460 lung cancer cells. For the first time, combined PANORAMA and fluorescence imaging enables quantitative analysis of microRNA-21 (miR-21) expression in sEVs to identify "cancer-suspicious" sub-population from liver cancer patient plasma in an unbiased fashion allowing both highly sensitive detection of individual sEVs and simultaneous molecular profiling. Collectively, NSSL enables uniform, high-performance plasmonic biosensing over large areas, providing a scalable and economical pathway for high-throughput, digital single-sEV analysis and translational liquid biopsy applications.

bioengineering↗

bioRxiv2025

Microfluidic Nano-Plasmonic Imaging Platform for Purification- and Label-Free Single Small Extracellular Vesicle Counting

Tumor-derived circulating small extracellular vesicles (sEVs) is a promising non-invasive biomarker for disease diagnosis. However, their quantitative detection remains challenging due to their small size and the complexity of blood plasma. Typically, sample preparation like purification is required. This study presents a purification-free approach using a microfluidic chip integrated with PANORAMA (Plasmonic nano-aperture label-free imaging) for label-free single sEV counting in plasma. CD63, CD9, and CD81 antibodies, specific biomarkers for most sEVs, are functionalized on AGNIS (arrayed gold nanodisks on invisible substrate) for selective capture. The automated microfluidic platform minimizes manual errors and allows precise programming of flow rates, directions, and media for optimization. Only 20 {micro}L of plasma is required, and the analysis is completed within 60 minutes. This platform shows great potential as a sensitive and efficient tool for detecting circulating sEVs without purification or labeling.

bioengineering↗

bioRxiv2020

Directed assembly and concentrating of micro/nanoparticles, cells, and vesicles via low-power near-infrared laser generated plasmonic microbubbles

Directed assembly and concentrating of micro- and nanoparticles via laser generated plasmonic microbubbles in a liquid environment is an emerging technology. For effective heating, visible light has been primarily employed in existing demonstrations. In this paper, we demonstrate a new plasmonic platform based on nanoporous gold disk (NPGD) array. Thanks to the highly tunable localized surface plasmon resonance of the NPGD array, microbubble of controlled size can be generated by near-infrared (NIR) light. Using NIR light provides several key advantages over visible light in less interference with standard microscopy and fluorescence imaging, preventing fluorescence photobleaching, less susceptible to absorption and scattering in turbid biological media, and much reduced photochemistry, phototoxicity and whatsoever. The large surface-to-volume ratio of NPGD further facilitates the heat transfer from these gold nanoheaters to the surroundings, achieving unprecedented low-power operation. While the microbubble is formed, the surrounding liquid circulates and direct microparticles randomly dispersed in the liquid to the bottom NPGD surface, yielding unique assemblies of microstructures. Such capability can also be employed in concentrating suspended colloidal nanoparticles at desirable sites and with preferred configuration, both enhancing the sensor performance. In addition to various micro- and nanoparticles, the plasmonic microbubbles are also shown to collect biological cells and nanovesicles. By using a spatial light modulator (SLM) to project the laser in arbitrary patterns, parallel assembly can be achieved to fabricate an array of clusters. These assemblies have been characterized using optical microscopy, scanning electron microscope, hyperspectral localized surface plasmon resonance imaging and hyperspectral Raman imaging.

bioengineering↗