MIFlowCyt-EV: a framework for standardized reporting of extracellular vesicle flow cytometry experiments

Joshua A Welsh; Edwin Van Der Pol; Ger J A Arkesteijn; Michel Bremer; Alain Brisson; Frank Coumans; Françoise Dignat-George; Erika Duggan; Ionita Ghiran; Bernd Giebel; André Görgens; An Hendrix; Romaric Lacroix; Joanne Lannigan; Sten F W M Libregts; Estefanía Lozano-Andrés; Aizea Morales-Kastresana; Stephane Robert; Leonie De Rond; Tobias Tertel; John Tigges; Olivier De Wever; Xiaomei Yan; Rienk Nieuwland; Marca H M Wauben; John P Nolan; Jennifer C Jones

doi:10.1080/20013078.2020.1713526

MIFlowCyt-EV: a framework for standardized reporting of extracellular vesicle flow cytometry experiments

J Extracell Vesicles. 2020 Feb 3;9(1):1713526. doi: 10.1080/20013078.2020.1713526. eCollection 2020.

Authors

Joshua A Welsh¹, Edwin Van Der Pol^{2

3

4}, Ger J A Arkesteijn⁵, Michel Bremer⁶, Alain Brisson⁷, Frank Coumans^{2

3

4}, Françoise Dignat-George^{8

9}, Erika Duggan¹⁰, Ionita Ghiran¹¹, Bernd Giebel⁶, André Görgens^{6

12

13}, An Hendrix¹⁴, Romaric Lacroix^{8

9}, Joanne Lannigan¹⁵, Sten F W M Libregts^{5

16}, Estefanía Lozano-Andrés⁵, Aizea Morales-Kastresana¹, Stephane Robert¹³, Leonie De Rond^{2

3

4}, Tobias Tertel⁶, John Tigges^{11

17}, Olivier De Wever¹⁴, Xiaomei Yan¹⁸, Rienk Nieuwland^{3

4}, Marca H M Wauben⁵, John P Nolan¹⁰, Jennifer C Jones¹

Affiliations

¹ Translational Nanobiology Section, Laboratory of Pathology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD, USA.
² Biomedical Engineering and Physics, Amsterdam UMC, University of Amsterdam, Amsterdam, The Netherlands.
³ Laboratory Experimental Clinical Chemistry, Amsterdam UMC, University of Amsterdam, Amsterdam, The Netherlands.
⁴ Vesicle Observation Center, Amsterdam UMC, University of Amsterdam, Amsterdam, The Netherlands.
⁵ Department of Biochemistry and Cell Biology, Faculty of Veterinary Medicine, Utrecht University, Utrecht, The Netherlands.
⁶ Institute for Transfusion Medicine, University Hospital Essen, University of Duisburg-Essen, Essen, Germany.
⁷ UMR-5248-CBMN, CNRS-University of Bordeaux-IPB, Pessac, France.
⁸ Center of Cardiovascular Research and Nutrition (C2VN) UMR-INSERM INRA 1263, Aix-Marseille Université, INSERM, Marseille, France.
⁹ Hematology and Vascular Biology Department, CHU La Conception, Assistance Publique-Hôpitaux de Marseille, Marseille, France.
¹⁰ Scintillon Institute, San Diego, CA, USA.
¹¹ Department of Medicine, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA, USA.
¹² Clinical Research Center, Department for Laboratory Medicine, Karolinska Institutet, Stockholm, Sweden.
¹³ Evox Therapeutics Ltd, Oxford, UK.
¹⁴ Laboratory of Experimental Cancer Research, Department of Human Structure and Repair, Ghent University Hospital, Ghent, Belgium.
¹⁵ Flow Cytometry Core, School of Medicine, University of Virginia, Charlottesville, VA, USA.
¹⁶ NIHR Cambridge BRC Cell Phenotyping Hub, Department of Medicine, University of Cambridge, Cambridge, UK.
¹⁷ Flow Cytometry Core, Beth Israel Deaconess Medical Center, Boston, MA, USA.
¹⁸ MOE Key Laboratory of Spectrochemical Analysis & Instrumentation, Key Laboratory for Chemical Biology of Fujian Province, Department of Chemical Biology, College of Chemistry and Chemical Engineering, Xiamen University, Xiamen, People's Republic of China.

Abstract

Extracellular vesicles (EVs) are small, heterogeneous and difficult to measure. Flow cytometry (FC) is a key technology for the measurement of individual particles, but its application to the analysis of EVs and other submicron particles has presented many challenges and has produced a number of controversial results, in part due to limitations of instrument detection, lack of robust methods and ambiguities in how data should be interpreted. These complications are exacerbated by the field's lack of a robust reporting framework, and many EV-FC manuscripts include incomplete descriptions of methods and results, contain artefacts stemming from an insufficient instrument sensitivity and inappropriate experimental design and lack appropriate calibration and standardization. To address these issues, a working group (WG) of EV-FC researchers from ISEV, ISAC and ISTH, worked together as an EV-FC WG and developed a consensus framework for the minimum information that should be provided regarding EV-FC. This framework incorporates the existing Minimum Information for Studies of EVs (MISEV) guidelines and Minimum Information about a FC experiment (MIFlowCyt) standard in an EV-FC-specific reporting framework (MIFlowCyt-EV) that supports reporting of critical information related to sample staining, EV detection and measurement and experimental design in manuscripts that report EV-FC data. MIFlowCyt-EV provides a structure for sharing EV-FC results, but it does not prescribe specific protocols, as there will continue to be rapid evolution of instruments and methods for the foreseeable future. MIFlowCyt-EV accommodates this evolution, while providing information needed to evaluate and compare different approaches. Because MIFlowCyt-EV will ensure consistency in the manner of reporting of EV-FC studies, over time we expect that adoption of MIFlowCyt-EV as a standard for reporting EV- FC studies will improve the ability to quantitatively compare results from different laboratories and to support the development of new instruments and assays for improved measurement of EVs.

Keywords: Extracellular vesicles; flow cytometry; framework; reporting; standardization.

Abstract

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