title: Reproducible extracellular vesicle size and concentration determination with tunable resistive pulse sensing.
abstract: BACKGROUND
  The size of extracellular vesicles (EVs) can be determined with a tunable resistive pulse sensor (TRPS). Because the sensing pore diameter varies from pore to pore, the minimum detectable diameter also varies. The aim of this study is to determine and improve the reproducibility of TRPS measurements.
  
  
  METHODS
  Experiments were performed with the qNano system (Izon) using beads and a standard urine vesicle sample. With a combination of voltage and stretch that yields a high blockade height, we investigate whether the minimum detected diameter is more reproducible when we configure the instrument targeting (a) fixed stretch and voltage, or (b) fixed blockade height.
  
  
  RESULTS
  Daily measurements with a fixed stretch and voltage (n=102) on a standard urine sample show a minimum detected vesicle diameter of 128±19 nm [mean±standard deviation; coefficient of variation (CV) 14.8%]. The vesicle concentration was 2.4·109±3.8·109 vesicles/mL (range 1.4·108-1.8·1010). When we compared setting a fixed stretch and voltage to setting a fixed blockade height on 3 different pores, we found a minimum detected vesicle diameter of 118 nm (CV 15.5%, stretch), and 123 nm (CV 4.5%, blockade height). The detected vesicle concentration was 3.2-8.2·108 vesicles/mL with fixed stretch and 6.4-7.8·108 vesicles/mL with fixed blockade height. Summary/conclusion: Pore-to-pore variability is the cause of the variation in minimum detected size when setting a fixed stretch and voltage. The reproducibility of the minimum detectable diameter is much improved by setting a fixed blockade height.
authors.affiliation: Department of Clinical Chemistry, Academic Medical Centre, University of Amsterdam, Amsterdam, The Netherlands; Department of Biomedical Engineering & Physics, Academic Medical Centre, University of Amsterdam, Amsterdam, The Netherlands; f.a.coumans@amc.uva.nl.
authors.forename: Frank A W
authors.initials: FA
authors.lastname: Coumans
authors.affiliation: Department of Clinical Chemistry, Academic Medical Centre, University of Amsterdam, Amsterdam, The Netherlands; Department of Biomedical Engineering & Physics, Academic Medical Centre, University of Amsterdam, Amsterdam, The Netherlands.
authors.forename: Edwin
authors.initials: E
authors.lastname: van der Pol
authors.affiliation: Department of Clinical Chemistry, Academic Medical Centre, University of Amsterdam, Amsterdam, The Netherlands.
authors.forename: Anita N
authors.initials: AN
authors.lastname: Böing
authors.affiliation: Department of Clinical Chemistry, Academic Medical Centre, University of Amsterdam, Amsterdam, The Netherlands.
authors.forename: Najat
authors.initials: N
authors.lastname: Hajji
authors.affiliation: Department of Clinical Chemistry, Academic Medical Centre, University of Amsterdam, Amsterdam, The Netherlands.
authors.forename: Guus
authors.initials: G
authors.lastname: Sturk
authors.affiliation: Department of Biomedical Engineering & Physics, Academic Medical Centre, University of Amsterdam, Amsterdam, The Netherlands.
authors.forename: Ton G
authors.initials: TG
authors.lastname: van Leeuwen
authors.affiliation: Department of Clinical Chemistry, Academic Medical Centre, University of Amsterdam, Amsterdam, The Netherlands.
authors.forename: Rienk
authors.initials: R
authors.lastname: Nieuwland
keywords: exosomes; extracellular vesicles; microparticles; nanoparticles; resistive pulse sensing; size determination