Reliable measurements of extracellular vesicles by clinical flow cytometry

Martine Kuiper, Arthur van de Nes, Rienk Nieuwland, Zoltan Varga, Edwin van der Pol

Research output: Contribution to journalArticlepeer-review


Extracellular vesicles (EVs) are cell-derived particles with a phospholipid membrane present in all body fluids. Because EV properties change in health and disease, EVs have excellent potential to become biomarkers for diagnosis, prognosis, or monitoring of disease. The only technique capable of detecting, sizing, and phenotyping a million of EVs within minutes is (clinical) flow cytometry. A flow cytometer measures light scattering and fluorescence signals of single EVs. Although these signals contain valuable information about the presence and composition of EVs, the signals are expressed in arbitrary units, which make the comparison of measurement results impossible between instruments and laboratories. Additionally, unintended and undocumented variations in the source, preparation, and analysis of the sample lead to orders of magnitude variations in the measured EV concentrations. Here, we will explain the basics, challenges, and common misconceptions of EV flow cytometry. In addition, we provide an overview of recent standardization initiatives, which are a prerequisite for comparison of clinical data and thus for clinical biomarker exploration of EVs.

Original languageEnglish (US)
JournalAmerican Journal of Reproductive Immunology
StateAccepted/In press - 2020

Bibliographical note

Funding Information:
This work is part of the project “METVES II” and received funding from the EMPIR program co‐financed by the Participating States and from the European Union's Horizon 2020 research and innovation program. E. van der Pol acknowledges funding from the Netherlands Organisation for Scientific Research‐Domain Applied and Engineering Sciences (NWO‐TTW), research program VENI 15924. We thank Aleksandra Gąsecka for providing the data underlying Figure 3 .


  • calibration
  • data interpretation
  • extracellular vesicles
  • flow cytometry
  • standardization

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