Optimized AF4 combined with density cushion ultracentrifugation enables profiling of high-purity human blood extracellular vesicles.

Publisher: Wiley Country of Publication: United States NLM ID: 101610479 Publication Model: Print Cited Medium: Internet ISSN: 2001-3078 (Electronic) Linking ISSN: 20013078 NLM ISO Abbreviation: J Extracell Vesicles Subsets: MEDLINE

Publication: 2020- : [Hoboken, NJ] : Wiley
Original Publication: Järfälla : Co-Action Pub.

Extracellular Vesicles*/metabolism ; Extracellular Vesicles*/chemistry ; Ultracentrifugation*/methods; Humans ; Proteomics/methods ; MicroRNAs/blood ; Fractionation, Field Flow/methods ; Biomarkers/blood ; Liquid Biopsy/methods ; Centrifugation, Density Gradient/methods

Extracellular vesicles (EVs) have emerged as a promising tool for clinical liquid biopsy. However, the identification of EVs derived from blood samples is hindered by the presence of abundant plasma proteins, which impairs the downstream biochemical analysis of EV-associated proteins and nucleic acids. Here, we employed optimized asymmetric flow field-flow fractionation (AF4) combined with density cushion ultracentrifugation (UC) to obtain high-purity and intact EVs with very low lipoprotein contamination from human plasma and serum. Further proteomic analysis revealed more than 1000 EV-associated proteins, a large proportion of which has not been previously reported. Specifically, we found that cell-line-derived EV markers are incompatible with the identification of plasma-EVs and proposed that the proteins MYCT1, TSPAN14, MPIG6B and MYADM, as well as the traditional EV markers CD63 and CD147, are plasma-EV markers. Benefiting from the high-purity of EVs, we conducted comprehensive miRNA profiling of plasma EVs and nanosized particles (NPs), as well as compared plasma- and serum-derived EVs, which provides a valuable resource for the EV research community. Overall, our findings provide a comprehensive assessment of human blood EVs as a basis for clinical biopsy applications.
(© 2024 The Author(s). Journal of Extracellular Vesicles published by Wiley Periodicals LLC on behalf of International Society for Extracellular Vesicles.)

Anal Chim Acta. 2020 Aug 29;1127:234-245. (PMID: 32800129)
Curr Biol. 2009 Dec 1;19(22):1875-85. (PMID: 19896381)
Nucleic Acids Res. 2015 Apr 20;43(7):e47. (PMID: 25605792)
J Extracell Vesicles. 2023 May;12(5):e12315. (PMID: 37202906)
Nat Cell Biol. 2021 Dec;23(12):1240-1254. (PMID: 34887515)
J Extracell Vesicles. 2022 May;11(5):e12213. (PMID: 35524458)
iScience. 2022 Mar 01;25(4):104012. (PMID: 35340435)
Anal Chem. 2022 Oct 18;94(41):14083-14091. (PMID: 36191238)
Proteomics. 2013 Nov;13(22):3354-64. (PMID: 24115447)
Research (Wash D C). 2023 Apr 13;6:0114. (PMID: 37223486)
Bioinformatics. 2009 Apr 15;25(8):1091-3. (PMID: 19237447)
J Extracell Vesicles. 2014 Jul 08;3:. (PMID: 25018865)
Genome Biol. 2009;10(3):R25. (PMID: 19261174)
Signal Transduct Target Ther. 2020 Aug 3;5(1):144. (PMID: 32747657)
Nucleic Acids Res. 2022 Jan 7;50(D1):D1522-D1527. (PMID: 34871441)
Nucleic Acids Res. 2014;42(17):e133. (PMID: 25063298)
Cell. 2017 Jan 12;168(1-2):252-263.e14. (PMID: 28017328)
Nucleic Acids Res. 2019 Jan 8;47(D1):D1211-D1217. (PMID: 30252093)
OMICS. 2012 May;16(5):284-7. (PMID: 22455463)
Bioinformatics. 2014 Aug 1;30(15):2114-20. (PMID: 24695404)
Genome Biol. 2014;15(12):550. (PMID: 25516281)
J Extracell Vesicles. 2024 Feb;13(2):e12404. (PMID: 38326288)
Nucleic Acids Res. 2012 Jan;40(1):37-52. (PMID: 21911355)
Nat Protoc. 2019 Apr;14(4):1027-1053. (PMID: 30833697)
Nat Cell Biol. 2018 Mar;20(3):332-343. (PMID: 29459780)
Cell. 2019 Apr 4;177(2):428-445.e18. (PMID: 30951670)
Anal Chim Acta. 2020 Aug 8;1124:137-145. (PMID: 32534666)
Cell. 2016 Mar 10;164(6):1226-1232. (PMID: 26967288)
Int J Biol Sci. 2021 Jan 1;17(1):163-177. (PMID: 33390841)
J Extracell Vesicles. 2014 Dec 22;3:26913. (PMID: 25536934)
Nat Methods. 2017 Feb 28;14(3):228-232. (PMID: 28245209)
Nature. 2022 Jan;601(7893):446-451. (PMID: 34937935)
Nucleic Acids Res. 2003 Jan 1;31(1):439-41. (PMID: 12520045)
J Intensive Care. 2015 Jan 07;3(1):2. (PMID: 25705427)
J Extracell Vesicles. 2022 Sep;11(9):e12266. (PMID: 36124834)
Clin Exp Med. 2023 Oct;23(6):2867-2875. (PMID: 36826611)
Nat Rev Mol Cell Biol. 2018 Apr;19(4):213-228. (PMID: 29339798)
Genome Res. 2003 Nov;13(11):2498-504. (PMID: 14597658)
EMBO J. 2019 May 2;38(9):. (PMID: 30975688)
Front Bioeng Biotechnol. 2022 Oct 17;10:948959. (PMID: 36324901)
Cancer Cell. 2016 Dec 12;30(6):836-848. (PMID: 27960084)
J Proteome Res. 2008 Aug;7(8):3475-80. (PMID: 18570454)
PLoS One. 2020 Aug 19;15(8):e0236439. (PMID: 32813744)
Bioinformation. 2007 May 20;2(1):5-7. (PMID: 18084642)
Cell Mol Life Sci. 2018 Aug;75(15):2873-2886. (PMID: 29441425)
BMC Bioinformatics. 2008 Dec 29;9:559. (PMID: 19114008)
STAR Protoc. 2020 Dec 22;2(1):100225. (PMID: 33786456)
Anal Bioanal Chem. 2017 Apr;409(10):2501-2518. (PMID: 28116495)
Int J Mol Sci. 2019 Dec 25;21(1):. (PMID: 31881680)
Methods Enzymol. 2020;645:155-180. (PMID: 33565970)
Nature. 2015 Jul 9;523(7559):177-82. (PMID: 26106858)
Anal Bioanal Chem. 2019 Jan;411(3):777-786. (PMID: 30470915)
Theranostics. 2020 Feb 19;10(8):3684-3707. (PMID: 32206116)
J Extracell Vesicles. 2020 Dec;10(2):e12044. (PMID: 33489012)
Cell. 2020 Aug 20;182(4):1044-1061.e18. (PMID: 32795414)
Theranostics. 2017 Jan 26;7(3):789-804. (PMID: 28255367)
Cancer Lett. 2022 Sep 1;543:215796. (PMID: 35728740)
Nucleic Acids Res. 2006 Jan 1;34(Database issue):D140-4. (PMID: 16381832)
Circ Res. 2017 Sep 29;121(8):920-922. (PMID: 28963190)
Sci Rep. 2016 Apr 18;6:24316. (PMID: 27087061)
J Extracell Vesicles. 2023 Jul;12(7):e12346. (PMID: 37422692)
Anal Chim Acta. 2009 Mar 9;635(2):132-43. (PMID: 19216870)
EMBO J. 2022 Sep 15;41(18):e109288. (PMID: 36052513)
J Extracell Vesicles. 2016 Oct 31;5:32945. (PMID: 27802845)
Nat Cell Biol. 2021 Jun;23(6):631-641. (PMID: 34108659)
Anal Chem. 2015 Sep 15;87(18):9225-33. (PMID: 26291637)

ZL-SRPG2201901 R&D Program of Guangzhou Laboratory; 2021QN02Y939 Guangdong Province High-level Talent Youth Project; 82200884 National Natural Science Foundation of China

Keywords: AF4; extracellular vesicles; human blood; liquid biopsy; plasma and serum

0 (MicroRNAs)
0 (Biomarkers)

Date Created: 20240713 Date Completed: 20240713 Latest Revision: 20240718

20240718

PMC11245685

10.1002/jev2.12470

39001700