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Calibration of Flow Cytometers Enables Reproducible Measurements of Extracellular Vesicle Concentrations and Reference Range Establishment

by Bettin, Britta A. , Antovic, Jovan P. , Robert, Stéphane , Nelissen, Inge , Carney, Randy P. , Ajzenberg, Nadine , Norris, Philip J. , Jones, Jennifer C. , Dignat‐George, Françoise , Menezes, Erika G. Marques , Nieuwland, Rienk , Berckmans, Pascale , Mizenko, Rachel R. , Nolan, John , Tang, Vera A. , Varga, Zoltán , Lucien, Fabrice , Gollwitzer, Christian , Kim, Yohan , Giebel, Bernd , Lacroix, Romaric , Meyer, Annette , Siljander, Pia R.‐M. , Welsh, Joshua A. , Leeuwen, Ton G. , Maaninka, Katariina , Falkena, Kim , Tertel, Tobias , Broeck, Tina Van Den , Poudel, Sumeet , Lannigan, Joanne , Li, Bo , Wang, Lili , Pink, Desmond , Buzas, Edit I. , Cook, Sean , Faille, Dorothee , Pol, Edwin

in Adult / Biomarkers - blood / Blood / Blood levels / blood plasma / Blood Platelets - metabolism / Body fluids / Calibration / Erythrocytes - metabolism / Extracellular vesicles / Extracellular Vesicles - metabolism / Female / Flow cytometry / Flow Cytometry - methods / Flow Cytometry - standards / Hemoglobin / Humans / International organizations / Leukocytes - metabolism / Life Sciences / Light / Male / Medical laboratories / Middle Aged / Plasma / reference ranges / Reference Values / reproducibility / Reproducibility of Results / Standardization

2025

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Calibration of Flow Cytometers Enables Reproducible Measurements of Extracellular Vesicle Concentrations and Reference Range Establishment

2025

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Calibration of Flow Cytometers Enables Reproducible Measurements of Extracellular Vesicle Concentrations and Reference Range Establishment

2025

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Journal Article

Calibration of Flow Cytometers Enables Reproducible Measurements of Extracellular Vesicle Concentrations and Reference Range Establishment

Bettin, Britta A.,

Antovic, Jovan P.,

Robert, Stéphane,

Nelissen, Inge,

Carney, Randy P.,

Ajzenberg, Nadine,

Norris, Philip J.,

Jones, Jennifer C.,

Dignat‐George, Françoise,

Menezes, Erika G. Marques,

Nieuwland, Rienk,

Berckmans, Pascale,

Mizenko, Rachel R.,

Nolan, John,

Tang, Vera A.,

Varga, Zoltán,

Lucien, Fabrice,

Gollwitzer, Christian,

Kim, Yohan,

Giebel, Bernd,

Lacroix, Romaric,

Meyer, Annette,

Siljander, Pia R.‐M.,

Welsh, Joshua A.,

Leeuwen, Ton G.,

Maaninka, Katariina,

Falkena, Kim,

Tertel, Tobias,

Broeck, Tina Van Den,

Poudel, Sumeet,

Lannigan, Joanne,

Li, Bo,

Wang, Lili,

Pink, Desmond,

Buzas, Edit I.,

Cook, Sean,

Faille, Dorothee,

Pol, Edwin

2025

Overview

The concentration of cells is a key component of modern blood tests. Given the biomarker potential of extracellular vesicles (EVs) in blood, we aimed to establish reference ranges for blood cell‐derived EVs using flow cytometry. To address the orders‐of‐magnitude variability in reported EV concentrations between different flow cytometers (FCMs), we first validated a calibration methodology to enable reproducible EV concentration measurements. The methodology was evaluated in an interlaboratory comparison study and shows that calibration reduces the median absolute deviation of EV concentrations measured on 25 different FCMs from 67 % to 25 %–31 %. The calibration methodology was then used to determine reference ranges of erythrocyte‐, leukocyte‐, and platelet‐derived EVs in human blood plasma in a cohort of healthy individuals (n = 224). This study demonstrates that calibration enables comparable concentration measurements of blood cell‐derived EVs, thereby bringing EVs one step closer to clinical applications.

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Publisher

John Wiley & Sons, Inc,Taylor & Francis,Wiley

Subject

/ Blood

/ Blood Platelets - metabolism

/ Body fluids