TY - JOUR
T1 - Inter-laboratory multiplex bead-based surface protein profiling of MSC-derived EV preparations identifies MSC-EV surface marker signatures
AU - Nguyen, Vivian V.T.
AU - Welsh, Joshua A.
AU - Tertel, Tobias
AU - Choo, Andre
AU - van de Wakker, Simonides I.
AU - Defourny, Kyra A.Y.
AU - Giebel, Bernd
AU - Vader, Pieter
AU - Padmanabhan, Jayanthi
AU - Lim, Sai Kiang
AU - Nolte-'t Hoen, Esther N.M.
AU - Verhaar, Marianne C.
AU - Bostancioglu, R. Beklem
AU - Zickler, Antje M.
AU - Hong, Jia Mei
AU - Jones, Jennifer C.
AU - EL Andaloussi, Samir
AU - van Balkom, Bas W.M.
AU - Görgens, André
N1 - Publisher Copyright:
© 2024 The Author(s). Journal of Extracellular Vesicles published by Wiley Periodicals LLC on behalf of International Society for Extracellular Vesicles.
PY - 2024/6
Y1 - 2024/6
N2 - Mesenchymal stromal cells (MSCs) are promising regenerative therapeutics that primarily exert their effects through secreted extracellular vesicles (EVs). These EVs – being small and non-living – are easier to handle and possess advantages over cellular products. Consequently, the therapeutic potential of MSC-EVs is increasingly investigated. However, due to variations in MSC-EV manufacturing strategies, MSC-EV products should be considered as highly diverse. Moreover, the diverse array of EV characterisation technologies used for MSC-EV characterisation further complicates reliable interlaboratory comparisons of published data. Consequently, this study aimed to establish a common method that can easily be used by various MSC-EV researchers to characterise MSC-EV preparations to facilitate interlaboratory comparisons. To this end, we conducted a comprehensive inter-laboratory assessment using a novel multiplex bead-based EV flow cytometry assay panel. This assessment involved 11 different MSC-EV products from five laboratories with varying MSC sources, culture conditions, and EV preparation methods. Through this assay panel covering a range of mostly MSC-related markers, we identified a set of cell surface markers consistently positive (CD44, CD73 and CD105) or negative (CD11b, CD45 and CD197) on EVs of all explored MSC-EV preparations. Hierarchical clustering analysis revealed distinct surface marker profiles associated with specific preparation processes and laboratory conditions. We propose CD73, CD105 and CD44 as robust positive markers for minimally identifying MSC-derived EVs and CD11b, CD14, CD19, CD45 and CD79 as reliable negative markers. Additionally, we highlight the influence of culture medium components, particularly human platelet lysate, on EV surface marker profiles, underscoring the influence of culture conditions on resulting EV products. This standardisable approach for MSC-EV surface marker profiling offers a tool for routine characterisation of manufactured EV products in pre-clinical and clinical research, enhances the quality control of MSC-EV preparations, and hopefully paves the way for higher consistency and reproducibility in the emerging therapeutic MSC-EV field.
AB - Mesenchymal stromal cells (MSCs) are promising regenerative therapeutics that primarily exert their effects through secreted extracellular vesicles (EVs). These EVs – being small and non-living – are easier to handle and possess advantages over cellular products. Consequently, the therapeutic potential of MSC-EVs is increasingly investigated. However, due to variations in MSC-EV manufacturing strategies, MSC-EV products should be considered as highly diverse. Moreover, the diverse array of EV characterisation technologies used for MSC-EV characterisation further complicates reliable interlaboratory comparisons of published data. Consequently, this study aimed to establish a common method that can easily be used by various MSC-EV researchers to characterise MSC-EV preparations to facilitate interlaboratory comparisons. To this end, we conducted a comprehensive inter-laboratory assessment using a novel multiplex bead-based EV flow cytometry assay panel. This assessment involved 11 different MSC-EV products from five laboratories with varying MSC sources, culture conditions, and EV preparation methods. Through this assay panel covering a range of mostly MSC-related markers, we identified a set of cell surface markers consistently positive (CD44, CD73 and CD105) or negative (CD11b, CD45 and CD197) on EVs of all explored MSC-EV preparations. Hierarchical clustering analysis revealed distinct surface marker profiles associated with specific preparation processes and laboratory conditions. We propose CD73, CD105 and CD44 as robust positive markers for minimally identifying MSC-derived EVs and CD11b, CD14, CD19, CD45 and CD79 as reliable negative markers. Additionally, we highlight the influence of culture medium components, particularly human platelet lysate, on EV surface marker profiles, underscoring the influence of culture conditions on resulting EV products. This standardisable approach for MSC-EV surface marker profiling offers a tool for routine characterisation of manufactured EV products in pre-clinical and clinical research, enhances the quality control of MSC-EV preparations, and hopefully paves the way for higher consistency and reproducibility in the emerging therapeutic MSC-EV field.
KW - culture conditions
KW - exosomes
KW - extracellular vesicles
KW - inter-laboratory assessment
KW - mesenchymal stromal cells
KW - MSC
KW - MSC-marker
KW - quality control
KW - regenerative medicine
KW - standardisation
KW - surface markers
UR - http://www.scopus.com/inward/record.url?scp=85196092247&partnerID=8YFLogxK
U2 - 10.1002/jev2.12463
DO - 10.1002/jev2.12463
M3 - Article
AN - SCOPUS:85196092247
SN - 2001-3078
VL - 13
JO - Journal of Extracellular Vesicles
JF - Journal of Extracellular Vesicles
IS - 6
M1 - e12463
ER -