TY - JOUR
T1 - Chemically-Induced Lipoprotein Breakdown for Improved Extracellular Vesicle Purification
AU - Iannotta, Dalila
AU - Amruta, A.
AU - Lai, Andrew
AU - Nair, Soumyalekshmi
AU - Koifman, Na'ama
AU - Lappas, Martha
AU - Salomon, Carlos
AU - Wolfram, Joy
N1 - Funding Information:
The authors thank Lifeblood Red Cross Australia for donating plasma. This work used the Queensland node of the National Collaborative Research Infrastructure Strategy‐enabled Australian National Fabrication Facility (ANFF). The Malvern NanoSight NS300 was purchased by Professor Matt Trau and associates under The University of Queensland Major Equipment and Infrastructure grant (2016000652). This work was partially funded by The University of Queensland, Australia (J.W.). C.S. is supported by the Lion Medical Research Foundation (2015001964), the Medical Research Future Fund (MRF1199984 and GA187319), and the National Health and Medical Research Council (NHMRC 1195451). The content is solely the responsibility of the authors and does not necessarily represent the official views of the funding agencies. Figures 1,2A,C–F,3C,4A,E,6A,D–F and the graphical abstract were completely or partially made in BioRender–biorender.com.
Publisher Copyright:
© 2023 The Authors. Small published by Wiley-VCH GmbH.
PY - 2024/5/2
Y1 - 2024/5/2
N2 - Extracellular vesicles (EVs) are nanosized biomolecular packages involved in intercellular communication. EVs are released by all cells, making them broadly applicable as therapeutic, diagnostic, and mechanistic components in (patho)physiology. Sample purity is critical for correctly attributing observed effects to EVs and for maximizing therapeutic and diagnostic performance. Lipoprotein contaminants represent a major challenge for sample purity. Lipoproteins are approximately six orders of magnitude more abundant in the blood circulation and overlap in size, shape, and density with EVs. This study represents the first example of an EV purification method based on the chemically-induced breakdown of lipoproteins. Specifically, a styrene-maleic acid (SMA) copolymer is used to selectively breakdown lipoproteins, enabling subsequent size-based separation of the breakdown products from plasma EVs. The use of the polymer followed by tangential flow filtration or size-exclusion chromatography results in improved EV yield, preservation of EV morphology, increased EV markers, and reduced contaminant markers. SMA-based EV purification enables improved fluorescent labeling, reduces interactions with macrophages, and enhances accuracy, sensitivity, and specificity to detect EV biomarkers, indicating benefits for various downstream applications. In conclusion, SMA is a simple and effective method to improve the purity and yield of plasma-derived EVs, which favorably impacts downstream applications.
AB - Extracellular vesicles (EVs) are nanosized biomolecular packages involved in intercellular communication. EVs are released by all cells, making them broadly applicable as therapeutic, diagnostic, and mechanistic components in (patho)physiology. Sample purity is critical for correctly attributing observed effects to EVs and for maximizing therapeutic and diagnostic performance. Lipoprotein contaminants represent a major challenge for sample purity. Lipoproteins are approximately six orders of magnitude more abundant in the blood circulation and overlap in size, shape, and density with EVs. This study represents the first example of an EV purification method based on the chemically-induced breakdown of lipoproteins. Specifically, a styrene-maleic acid (SMA) copolymer is used to selectively breakdown lipoproteins, enabling subsequent size-based separation of the breakdown products from plasma EVs. The use of the polymer followed by tangential flow filtration or size-exclusion chromatography results in improved EV yield, preservation of EV morphology, increased EV markers, and reduced contaminant markers. SMA-based EV purification enables improved fluorescent labeling, reduces interactions with macrophages, and enhances accuracy, sensitivity, and specificity to detect EV biomarkers, indicating benefits for various downstream applications. In conclusion, SMA is a simple and effective method to improve the purity and yield of plasma-derived EVs, which favorably impacts downstream applications.
KW - contaminants
KW - exosome
KW - isolation
KW - lipoproteins
KW - microvesicle
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U2 - 10.1002/smll.202307240
DO - 10.1002/smll.202307240
M3 - Article
AN - SCOPUS:85179661912
SN - 1613-6810
VL - 20
JO - Small
JF - Small
IS - 18
M1 - 2307240
ER -