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. 2021 Mar 22;11:6489. doi: 10.1038/s41598-021-85833-y

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© The Author(s) 2021

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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GF-EV-mediated delivery of exogenous proteins into the human cells analyzed by flow cytometry and confocal microscopy. (A) Incubation of colon cancer HCT-116 cells with GF-EVs loaded with BSA-AF647 or HSP70-AF647 proteins for 1 h. (B) Incubation of HCT-116 cells with HSP70-AF647-loaded GF-EVs for 1 and 3 h. (C,D) Incubation of HCT-116 cells with loaded GF-EVs in comparison with the free HSP70-AF647 or BSA-AF647 proteins for 1 h. (E,F) Incubation of peripheral blood mononuclear cells (E, monocytes and F, lymphocytes) with loaded GF-EVs in comparison with the free HSP70-AF647 or BSA-AF647 proteins for 1 h. (G,H) The relative intensity of fluorescence of HCT-116 cells and monocytes incubated with loaded GF-EVs in comparison with the free HSP70-AF647 or BSA-AF647 proteins. N = 2. Data are presented as means ± SD. *, p < 0.05, **; p < 0.01; ***, p < 0.001 as estimated by one-way ANOVA followed by multiple comparisons Tukey’s post-hoc analysis. (I,J) Micrographs of DLD1 cells co-cultivated with GF-EVs loaded with HSP70-AF647 (I) or BSA-AF647 (J) proteins. Scale bars are 20 µm. (K) Accumulation of fluorescent protein delivered into the recipient DLD1 cells by GF-EVs. Video recording frames were captured every 30 min after adding of loaded GF-EVs to the cells.