Analytical challenges of extracellular vesicle detection: A comparison of different techniques.

The interest in extracellular vesicles (EVs) has grown exponentially over the last decade. Evolving evidence is demonstrating that these EVs are playing an important role in health and disease. They are involved in intercellular communication and have been shown to transfer proteins, lipids, and nucleic acids. This review focuses on the most commonly used techniques for detection of EVs, to include microparticles, 100-1,000 nm in size, and exosomes, 50-100 nm in size. Conventional flow cytometry is the most prevalent technique, but nanoparticle tracking analysis (NTA), dynamic light scattering (DLS), and resistive pulse sensing have also been used to detect EVs. The accurate measurement of these vesicles is challenged by size heterogeneity, low refractive index, and the lack of dynamic measurement range for most of the available technologies. Sample handling during the preanalytical phase can also affect the accuracy of measurements. Currently, there is not one single method which allows phenotyping, sizing, and enumerating the whole range of EVs and, therefore, providing all the necessary information to truly understand the biology of these particles. A combination of methods is probably needed which might also include electron and atomic force microscopy and full RNA, lipid, and protein profiling.