biostudies-literaturede Voogt WSDutch Research Council (NWO), Domain Applied and Engineering SciencesDutch Research Council (NWO)e70017https://www.ebi.ac.uk/biostudies/studies/S-EPMC11653091biostudies-literatureUnknown13(12)Intercellular communication via extracellular vesicles (EVs) has been identified as a vital component of a steadily expanding number of physiological and pathological processes. To accommodate these roles, EVs have highly heterogeneous molecular compositions. Given that surface molecules on EVs determine their interactions with their environment, EV functionality likely differs between subpopulations with varying surface compositions. However, it has been technically challenging to examine such functional heterogeneity due to a lack of non-destructive methods to separate EV subpopulations based on their surface markers. Here, we used the Design-of-Experiments (DoE) methodology to optimize a protocol, which we name 'EV-Elute', to elute intact EVs from commercially available Protein G-coatedJournal of extracellular vesiclesEV-Elute: A universal platform for the enrichment of functional surface marker-defined extracellular vesicle subpopulations.PMC1165309117296VI.Veni.192.174Frunt RPaspali Ide Jong OGMonica BFrancois JJJMTriesscheijn CSKooijmans SAATielemans Mde Voogt WSLeandro RMSeinen CWfalseEV-Elute: A universal platform for the enrichment of functional surface marker-defined extracellular vesicle subpopulations.Intercellular communication via extracellular vesicles (EVs) has been identified as a vital component of a steadily expanding number of physiological and pathological processes. To accommodate these roles, EVs have highly heterogeneous molecular compositions. Given that surface molecules on EVs determine their interactions with their environment, EV functionality likely differs between subpopulations with varying surface compositions. However, it has been technically challenging to examine such functional heterogeneity due to a lack of non-destructive methods to separate EV subpopulations based on their surface markers. Here, we used the Design-of-Experiments (DoE) methodology to optimize a protocol, which we name 'EV-Elute', to elute intact EVs from commercially available Protein G-coated2024-01-01T00:00:00Z2024 Dec2025-04-22T10:40:43.324Z2025-04-05T23:42:49.216ZS-EPMC116530913969211510.1002/jev2.70017