A standardized method to determine the concentration of extracellular vesicles using tunable resistive pulse sensing

Robert Vogel, Frank A.W. Coumans, Raluca G. Maltesen, Anita N. Böing, Katherine E. Bonnington, Marike L. Broekman, Murray F. Broom, Edit I. Buzás, Gunna Christiansen, Najat Hajji, Søren R. Kristensen, Meta J. Kuehn, Sigrid M. Lund, Sybren L.N. Maas, Rienk Nieuwland, Xabier Osteikoetxea, Rosalie Schnoor, Benjamin J. Scicluna, Mitch Shambrook, Jeroen de VrijStephen I. Mann, Andrew F. Hill, Shona Pedersen*

*Corresponding author for this work

Research output: Contribution to journalArticleAcademicpeer-review

139 Citations (Scopus)
17 Downloads (Pure)


Background: Understanding the pathogenic role of extracellular vesicles (EVs) in disease and their potential diagnostic and therapeutic utility is extremely reliant on in-depth quantification, measurement and identification of EV sub-populations. Quantification of EVs has presented several challenges, predominantly due to the small size of vesicles such as exosomes and the availability of various technologies to measure nanosized particles, each technology having its own limitations. Materials and Methods: A standardized methodology to measure the concentration of extracellular vesicles (EVs) has been developed and tested. The method is based on measuring the EVconcentration as a function of a defined size range. Blood plasma EVs are isolated and purified using size exclusion columns (qEV) and consecutively measured with tunable resistive pulse sensing (TRPS). Six independent research groups measured liposome and EV samples with the aim to evaluate the developed methodology. Each group measured identical samples using up to 5 nanopores with 3 repeat measurements per pore. Descriptive statistics and unsupervised multivariate data analysis with principal component analysis (PCA) were used to evaluate reproducibility across the groups and to explore and visualise possible patterns and outliers in EV and liposome data sets. Results: PCA revealed good reproducibility within and between laboratories, with few minor outlying samples. Measured mean liposome (not filtered with qEV) and EV (filtered with qEV) concentrations had coefficients of variance of 23.9% and 52.5%, respectively. The increased variance of the EV concentration measurements could be attributed to the use of qEVs and the polydisperse nature of EVs. Conclusion: The results of this study demonstrate the feasibility of this standardized methodology to facilitate comparable and reproducible EV concentration measurements.

Original languageEnglish
Article number31242
JournalJournal of Extracellular Vesicles
Issue number1
Early online date29 Sept 2016
Publication statusPublished - 2016
Externally publishedYes


  • Colloids
  • Concentration
  • Coulter counter
  • EV
  • Exosomes
  • Extracellular vesicles
  • Microparticles
  • Micropores
  • Nanoparticles
  • Nanopores
  • Resistive pulse sensing


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