Monocrystalline Labeling Enables Stable Plasmonic Enhancement for Isolation‐Free Extracellular Vesicle Analysis

Abstract

such applications due to their straightforward synthesis, optical properties, and the diverse array of surface chemistry modifications that can be used to functionalize them with factors that recognize specific biomolecules and thereby generate probes that can detect and quantify specific targets of interest. [2] Such NP probes are typically highly stable, the signal they produce is durable, and can be more intense than that produced by fluorescent dyes or quantum dots, yet can be read by inexpensive dark field microscopes available in most research and clinical laboratories. [3] Scattering signal corresponds to NP size, although the inverse relationship between NP signal and NP binding kinetics, can constrain the size range of NPs used in nanoprobe applications and their limits of detection. Alternate detection approaches that employ enzymatic reactions to enhance signal intensity (e.g., an electrochemical-chemical-enzymatic redox cycling reaction read by an electrochemical sensor [4] and conversion of enzyme substrate to an insoluble form read by a simple surface plasmonic resonance sensor [5] ) can be more sensitive but may need to be read in real-time, require sophisticated equipment and highly trained personnel, or depend upon close temperature control during transport, storage, and assay performance.The signal produced by NP probes are a function of their composition, geometry, and size but can be modified in situ by increasing the apparent size or surface composition of affinity bound NPs through specific NP growth, modification, or aggregation methods. Multiple methods have been developed to enhance biosensing characteristics of localized surface plasmonic resonance (LSPR) probes, but most of these involve tradeoffs between signal enhancement efficiency, target specificity, and assay complexity. For example, NPs with complex (e.g., nanostars) or hybrid compositions (e.g., gold/silica structures) can exhibit enhanced light scattering characteristics, but these material can be difficult to synthesis and manipulate, [6] and increasing NP size to improve scattering can negatively affect their binding kinetics. [7] However, chemical reactions employed to induce in situ NP growth or surface modifications after their binding to a biomarker target can disrupt probetarget interactions to produce signal loss or allow nonspecific deposition of NP material on other matrixes present the Sensitive detection of extracellular vesicles (EVs) as emerging biomarkers has shown great promises for disease diagnosis. Plasmonic metal nanostructures conjugated with molecules that bind specific biomarker targets are widely used for EVs sensing but involve tradeoffs between particle-sizedependent signal intensity and conjugation efficiency. One solution to this problem would be to induce nucleation on nanoparticles that have successfully bound a target biomarker to permit in situ nanoparticle growth for signal amplification, but approaches that are evaluated to date require harsh conditions or lack nucleation specificity, prohibi...

Progress in Isolation and Molecular Profiling of Small Extracellular Vesicles via Bead-Assisted Platforms

“…Rho J. et al developed a prototype micro-nuclear magnetic resonance (uNMR) system that allows to isolate and quantify microvesicles (MVs) in packed red blood cell (pRBC) units and label their subpopulations with target-specific magnetic nanoparticles [46]. In a recent study, Wang S. et al developed and tested a rapid Cu nanoshell-enhanced immunoassay (Cu-NEI) where in situ Cu growth allows to significantly enhance NP-induced signal intensity [76]. The group first tested and compared AuR with AuS enhancement by Cu nanoshell growth.…”

Colocalization of protein and microRNA markers reveals unique extracellular vesicle subpopulations for early cancer detection

“…Confocal fluorescence microscopy (CFM) or total internal reflection fluorescence microscopy has also come to the spot light of single EV analysis because of their relatively lower cost in instrumentation and less complex operation while offering direct visualization of the vesicles (28,36,37). As listed in table S1, the most state-of-the-art developments using fluorescence microscopy detected either proteins or nuclei acids on single EVs, confirming the presence of EV subpopulations bearing different phenotypic features and their premises in marking disease development (28,(38)(39)(40)(41)(42). Still, single EV analysis is very difficult, owing to their extremely small sizes and the low amounts of cargos enclosed in each EV.…”

“…Still, single EV analysis is very difficult, owing to their extremely small sizes and the low amounts of cargos enclosed in each EV. While impressive detection performance has been obtained with the pioneering developments, assay turnaround time, limit of detection (LOD), and sample consumption are yet to be improved to meet the needs in early detection and frequent disease monitoring (38)(39)(40)(41)(42). Single EV capture was only achieved with specially designed surface features fabricated on microfluidic devices or by controlling a large bead-to-EV molar ratio, which is not easy to do when testing unknown samples.…”

Choline Phosphate-Grafted Nanozymes as Universal Extracellular Vesicle Probes for Bladder Cancer Detection

“…Inorganic nanozymes are widely used in biomedical research because of their high catalytic efficiency and good stability, outperforming biological enzymes (such as horseradish peroxidase, HRP) for signal amplification in ELISA. , To endow nanozymes with specific recognition toward target proteins, the surfaces of nanozymes are often modified with antibodies or aptamers. − Considering that the catalytic activity of nanozymes depends on the surface’s active sites, these surface modification strategies may impair their enzymatic activity . Various kinds of antibodies or aptamers are involved in fabricating nanoprobes for multiplex profiling of EV protein biomarkers. − However, only a few nanoprobes can be recognized by EV membrane proteins due to the small size of EVs and low copy number of the target proteins (normally 1–5 per EV), − leading to false negative results.…”