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. 2020 Jun 1;10:8856. doi: 10.1038/s41598-020-65517-9

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© The Author(s) 2020

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 license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license 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 license, visit http://creativecommons.org/licenses/by/4.0/.

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pECs-EVs can be uptaken by spermatozoa in a co-culture system. pECs-EVs were labelled for 2 hours with Vybrant DiO dye, analyzed by flow cytometry (C) and live imaging (D) and allowed to release EVs for 48 hours. Physical parameters (A) and fluorescence (B) of unlabeled pECs were analyzed for additional 48 hours (see Materials and Methods section and Supplementary Figure 2). Uptake confirmation and efficiency were evaluated by flow cytometry (G) and live imaging snapshot (H). Physical parameters (E) and fluorescence (F) of spermatozoa before incubation with pECs are shown as controls. Sperm cells recovered from the co-culture demonstrated a diffused fluorescence both by flow cytometry (G) and live imaging snapshot (H).