Towards traceable size determination of extracellular vesicles

Abstract

BackgroundExtracellular vesicles (EVs) have clinical importance due to their roles in a wide range of biological processes. The detection and characterization of EVs are challenging because of their small size, low refractive index, and heterogeneity.MethodsIn this manuscript, the size distribution of an erythrocyte-derived EV sample is determined using state-of-the-art techniques such as nanoparticle tracking analysis, resistive pulse sensing, and electron microscopy, and novel techniques in the field, such as small-angle X-ray scattering (SAXS) and size exclusion chromatography coupled with dynamic light scattering detection.ResultsThe mode values of the size distributions of the studied erythrocyte EVs reported by the different methods show only small deviations around 130 nm, but there are differences in the widths of the size distributions.ConclusionSAXS is a promising technique with respect to traceability, as this technique was already applied for traceable size determination of solid nanoparticles in suspension. To reach the traceable measurement of EVs, monodisperse and highly concentrated samples are required.

Measurement of microvesicle levels in human blood using flow cytometry

“…The applicability and limitations of the different methods have been extensively reviewed (6,7), and comparisons made between methods using urinary microvesicles (42) and red cell microvesicles from outdated packed red cell units (43), both of which have high concentrations of relatively pure less variable populations than are found in whole blood.…”

“…Alternate protocols have been proposed for calibrating microvesicle size using polystyrene and silica beads of different sizes combined with computer calculations of equivalent microvesicle size (42,43). While fluorescence triggering has been shown to detect more microvesicles than light scattering detection on conventional flow cytometers, due to the variable nature of fluorescence labeling and triggering, no standardized method has been developed for calibrating microvesicle size limits using fluorescence measurements.…”

“…These methods are able to accurately separate lipid membrane vesicles from other particles in blood. Estimates of the peak size distribution for microvesicles in platelet free plasma using electron microscopy have ranged from 160 to 300 nm (Table 6) (26,30,43,53). Compared to other methods, electron microscopy is labor intensive and may underestimate the number of microvesicles due to sample preparation issues (42).…”

Exosomes: potential model for complement-stealth delivery systems

“…Other imaging technologies that have also been implemented for such use include micro nuclear magnetic resonance, and X-ray scattering (99)(100)(101)(102). Finding the origin of a population of exosomes extracted from a sample can be achieved by analyzing the specific marker antigens on the surface of the exosome.…”

Platelets and platelet extracellular vesicles as messengers in vascular inflammation

“…Further methods have been applied and adapted for the assessment of microvesicles. Electron microscopy (EM) and transmission electron microscopy (TEM) allows analyzing the morphology, size and presence of specific surface molecules [82]. Advances in electron microscopy might enable visualization of their 3 dimensional structures at high magnification [83].…”