Abstract
Extracellular vesicles (EVs) have been lauded as next generation medicines, but very few EV-based therapeutics have progressed to clinical use. Limited clinical translation is largely due to technical barriers that hamper our ability to mass-produce EVs, i.e. to isolate, purify and characterise them effectively. Technical limitations in comprehensive characterisation of EVs leads to unpredicted biological effects of EVs. Here, using a range of optical and non-optical techniques, we showed that the differences in molecular composition of EVs isolated using two isolation methods correlated with the differences in their biological function. Our results demonstrated that the isolation method determines the composition of isolated EVs at single and sub-population levels. Besides the composition, we measured for the first time the dry mass and predicted sedimentation of EVs. These parameters were shown to correlate well with the biological and functional effects of EVs on single cell and cell cultures. We anticipate that our multiscale characterisation approach will support fundamental understanding of EVs as well as elucidate the functional effects of EVs in in vitro and in vivo studies. Our findings and methodology will be pivotal for developing optimal isolation methods and establishing EVs as mainstream therapeutics and diagnostics. This innovative approach is applicable to a wide range of sectors including biopharma and biotechnology as well as to regulatory agencies.
Competing Interest Statement
The authors have declared no competing interest.
List of Abbreviations
- AFM
- Atomic Force Microscopy
- AFM-IR
- Atomic Force Microscope Infrared-Spectroscopy
- CMSC29
- chorionic mesenchymal stromal/stem cell line
- CEVs
- extracellular vesicles derived from chorionic mesenchymal stromal/stem cell
- DMSC23
- decidual mesenchymal stromal/stem cell line
- DEVs
- extracellular vesicles derived from decidual mesenchymal stromal/stem cell
- EVs
- extracellular vesicles
- HBSS(-)
- hanks’ balanced salt solution
- LPS
- lipopolysaccharide
- MSC
- mesenchymal stromal/stem cell
- PTA
- Particle Tracking Analysis
- nFCM
- Nano-flow Cytometry
- RMM
- Resonant Mass Measurement
- TRPS
- Tunable Resistive Pulse Sensing
- TFF
- tangential flow filtration