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. 2022 Apr 5;12:5651. doi: 10.1038/s41598-022-09627-6

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© The Author(s) 2022

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.

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Characterization and quantification of isolated EVs. (a) Flow cytometric analysis was performed using magnetic beads coated with CD9 EV marker. These coated beads were incubated with a PE-conjugated CD81 antibody. Analaysis demonstrated as dot plot. FSC represents forward scatter and SSC represents size vs. side scatter. P1 represents gate on the doublet discrimination according to FSC-H vs FSC-A. (b) Size distribution and quantification of isolated extracellular vesicles were analyzed using qNanoGold. A polyurethane nanopore rated for particles < 100 Npm (NP100-, Izon Science, UK) was used. The measured mean diameter of EVS was approximately 100 nm. (c) Western blot was performed to detect EV proteins and cellular contaminants in isolated EVs. Tsg101 was used as EV markers and endoplasmic reticulum marker Calnexin was used as a negative control. (d) EVs isolated from untreated H9c2 (Dil-Control-EV) and ticagrelor treated H9c2 (Dil-Tica-EV) stained with Dil dye. The uptake of DiI-labeled EVs by H9c2 cells was visualized by confocal microscopy. The cells incubated with Dil-labelled PBS are the negative control group. (e) EVs isolated from control and ticagrelor treated H9c2 cells were visualized by transmission electron microscopy (TEM).