Comparing digital detection platforms in high sensitivity immune‐phenotyping of extracellular vesicles

Abstract

Despite their clinical potential, Extracellular Vesicles (EVs) struggle to take the scene as a preeminent source of biomarkers in liquid biopsy. Limitations in the use of EVs origin from their inherent complexity and heterogeneity and from the sensitivity demand in detecting low to very low abundant disease-specific sub-populations. Such need can be met by digital detection, namely capable to reach the single-molecule sensitivity. Here we set to compare, side by side, two digital detection platforms that have recently gained increasing importance in the field of EVs. The platforms, both commercially available, are based on the principles of the Single Particle Interferometric Reflectance Imaging Sensing (SP-IRIS) and the Single Molecule Array technology (SiMoA) respectively. Sensitivity in immune-phenotyping of a well characterized EV sample is reported, discussing possible applicative implications and rationales for alternative or complementary use of the two platforms in biomarker discovery or validation.

Ultrasensitive and High‐Resolution Protein Spatially Decoding Framework for Tumor Extracellular Vesicles

“…Notably, the utilization of Simoa for EV protein analysis in cancer diagnostics is still in its infancy, with only five previous studies identified. [ 40–44 ]…”

“…Numerous EV analytical technologies have been previously published for the analysis of EV-associated proteins. [6,11,[39][40][41][42][43][44][45][46][47][48][49][50] Single-EV analysis (sEVA) employed a fluorescence imaging technique to determine the percentage of EVs positive for specific proteins. [6,50] This approach required high-resolution microscopy and the establishment of a threshold for considering vesicular fluorescence events as positive.…”

“…Notably, the utilization of Simoa for EV protein analysis in cancer diagnostics is still in its infancy, with only five previous studies identified. [40][41][42][43][44] Our work achieved three key advancements compared to these previous studies. [40][41][42][43][44] First, the eSimoa assays exhibited superior sensitivity compared to their respective assays.…”

“…[40][41][42][43][44] Our work achieved three key advancements compared to these previous studies. [40][41][42][43][44] First, the eSimoa assays exhibited superior sensitivity compared to their respective assays. Second, the eSimoa measured both EV surface and luminal proteins, whereas the previous studies were predominantly focused on surface proteins such as CD9, CD63, CD81, or PD-L1.…”

A multifarious platform for extracellular vesicles analysis

“…Extracellular Vesicles (EVs) are a highly heterogeneous population of biogenic nanoparticles enclosed in a lipid bilayer, released by almost all cell types, and involved in intercellular communication by transferring cellular biomolecules, among which proteins and nucleic acids. As EVs molecular compositions reflects the cellular source, in the health field they have arisen as potential biomarkers for liquid biopsies 1 . Moreover, thanks to their physical structure, the possibility of biotechnological engineering makes them also innovative therapeutic agents 2,3 .…”