Reproducibility of extracellular vesicle research

Cells release membrane-delimited particles into the environment. These particles are called “extracellular vesicles” (EVs), and EVs are present in fluids contacting cells, including body fluids and conditioned culture media. Because EVs change and contribute to health and disease, EVs have become a hot topic. From the thousands of papers now published on EVs annually, one easily gets the impression that EVs provide biomarkers for all diseases, and that EVs are carriers of all relevant biomolecules and are omnipotent therapeutics. At the same time, EVs are heterogeneous, elusive and difficult to study due to their physical properties and the complex composition of their environment.

This overview addresses the current challenges encountered when working with EVs, and how we envision that most of these challenges will be overcome in the near future. Right now, an infrastructure is being developed to improve the reproducibility of EV measurement results. This infrastructure comprises expert task forces of the International Society of Extracellular Vesicles (ISEV) developing guidelines and recommendations, instrument calibration, standardized and transparent reporting, and education. Altogether, these developments will support the credibility of EV research by introducing robust reproducibility, which is a prerequisite for understanding their biological significance and biomarker potential.

Towards reproducible measurements of extracellular vesicles

The current lack of reproducibility in extracellular vesicle (EV) research hinders progress in understanding their biological role and theranostic applicability. Isolation and analyses are hampered by the physical and biochemical heterogeneity of EVs, the complex composition of tissues and biofluids containing the EVs, unidentified variables that affect the presence and function of EVs in a biospecimen, and the lack of both instrument calibration and standardized reporting of methods and results. In the near future, based on ongoing developments, reproducibility will be greatly improved by guidelines for collection, handling and storage of biofluids (pre-analytics), minimal development of novel separation and characterization methods, instrument calibration, requirements for systematic reporting of relevant biospecimen variables, methods and results, defining minimal requirements relevant to each step, and education effort. Comparable and reproducible results will then enable multicenter studies required for biomarker discovery and clinical utility.

Nieuwland R, Siljander PR, Falcón-Pérez JM, Witwer KW. (2022) Reproducibility of extracellular vesicle research. Eur J Cell Biol 101(3):151226. [article]

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