title: Ultrafiltration with size-exclusion liquid chromatography for high yield isolation of extracellular vesicles preserving intact biophysical and functional properties.
abstract: Extracellular vesicles (EVs) are natural nanoparticles that mediate intercellular transfer of RNA and proteins and are of great medical interest; serving as novel biomarkers and potential therapeutic agents. However, there is little consensus on the most appropriate method to isolate high-yield and high-purity EVs from various biological fluids. Here, we describe a systematic comparison between two protocols for EV purification: ultrafiltration with subsequent liquid chromatography (UF-LC) and differential ultracentrifugation (UC). A significantly higher EV yield resulted from UF-LC as compared to UC, without affecting vesicle protein composition. Importantly, we provide novel evidence that, in contrast to UC-purified EVs, the biophysical properties of UF-LC-purified EVs are preserved, leading to a different in vivo biodistribution, with less accumulation in lungs. Finally, we show that UF-LC is scalable and adaptable for EV isolation from complex media types such as stem cell media, which is of huge significance for future clinical applications involving EVs.
  
  
  BACKGROUND
  Recent evidence suggests extracellular vesicles (EVs) as another route of cellular communication. These EVs may be utilized for future therapeutics. In this article, the authors compared ultrafiltration with size-exclusion liquid chromatography (UF-LC) and ultra-centrifugation (UC) for EV recovery.
authors: Department of Laboratory Medicine, Karolinska Institutet, Stockholm, Sweden.
authors: Joel Z
authors: JZ
authors: Nordin
authors: Department of Physiology, Anatomy and Genetics, University of Oxford, Oxford, United Kingdom.
authors: Yi
authors: Y
authors: Lee
authors: Department of Physiology, Anatomy and Genetics, University of Oxford, Oxford, United Kingdom.
authors: Pieter
authors: P
authors: Vader
authors: Department of Physiology, Anatomy and Genetics, University of Oxford, Oxford, United Kingdom; Institute of Technology, University of Tartu, Tartu, Estonia.
authors: Imre
authors: I
authors: Mäger
authors: Cancer Proteomics Mass Spectrometry, Science for Life Laboratory, Department of Oncology-Pathology, Karolinska Institutet, Stockholm, Sweden.
authors: Henrik J
authors: HJ
authors: Johansson
authors: Novartis Institutes for Biomedical Research, Basel, Switzerland.
authors: Wolf
authors: W
authors: Heusermann
authors: Department of Laboratory Medicine, Karolinska Institutet, Stockholm, Sweden.
authors: Oscar P B
authors: OP
authors: Wiklander
authors: Department of Neurochemistry, Stockholm University, Stockholm, Sweden.
authors: Mattias
authors: M
authors: Hällbrink
authors: Molecular Engineering Laboratory, Proteos, Agency for Science, Technology and Research (A*STAR), Singapore.
authors: Yiqi
authors: Y
authors: Seow
authors: Department of Medicine Solna, Translational Immunology Unit, Karolinska Institutet and University Hospital, Stockholm, Sweden.
authors: Jarred J
authors: JJ
authors: Bultema
authors: Umeå Centre for Molecular Medicine, Umeå University, Umeå, Sweden.
authors: Jonathan
authors: J
authors: Gilthorpe
authors: Sir William Dunn School of Pathology, University of Oxford, Oxford, United Kingdom.
authors: Tim
authors: T
authors: Davies
authors: Sir William Dunn School of Pathology, University of Oxford, Oxford, United Kingdom.
authors: Paul J
authors: PJ
authors: Fairchild
authors: Department of Medicine Solna, Translational Immunology Unit, Karolinska Institutet and University Hospital, Stockholm, Sweden.
authors: Susanne
authors: S
authors: Gabrielsson
authors: Novartis Institutes for Biomedical Research, Basel, Switzerland.
authors: Nicole C
authors: NC
authors: Meisner-Kober
authors: Cancer Proteomics Mass Spectrometry, Science for Life Laboratory, Department of Oncology-Pathology, Karolinska Institutet, Stockholm, Sweden.
authors: Janne
authors: J
authors: Lehtiö
authors: Department of Laboratory Medicine, Karolinska Institutet, Stockholm, Sweden.
authors: C I Edvard
authors: CI
authors: Smith
authors: Department of Physiology, Anatomy and Genetics, University of Oxford, Oxford, United Kingdom. Electronic address: matthew.wood@dpag.ox.ac.uk.
authors: Matthew J A
authors: MJ
authors: Wood
authors: Department of Laboratory Medicine, Karolinska Institutet, Stockholm, Sweden; Department of Physiology, Anatomy and Genetics, University of Oxford, Oxford, United Kingdom. Electronic address: Samir.El-Andaloussi@ki.se.
authors: Samir
authors: S
authors: El Andaloussi
mesh_terms: D055252:Cell-Derived Microparticles / Q000737:chemistry* / Q000648:ultrastructure*; D002850:Chromatography, Gel; D057809:HEK293 Cells; D006801:Humans; D014462:Ultrafiltration
keywords: Biophysical properties; extracellular vesicles; size-exclusion liquid chromatography; ultracentrifugation; ultrafiltration