Fig. 2: Development of the VFIC system to further improve intracellular protein delivery by EVs. | Nature Communications

Fig. 2: Development of the VFIC system to further improve intracellular protein delivery by EVs.

From: Engineering of extracellular vesicles for efficient intracellular delivery of multimodal therapeutics including genome editors

Fig. 2

a VSV-G+ and CD63 + EV concentrations as determined by single-vesicle flow cytometry after transfection with VSV-G-mNG alone or VSV-G-mNG and CD63 together. b Constructs generated for developing the VFIC system. The last construct was generated as a negative control where Cre was replaced with the bacteriophage protein MS249. c EV dose-dependent recombination in B16F10-TL cells mediated by VSV-G-Foldon-Intein-Cre and VSV-G-Intein-Cre EVs as evaluated by flow cytometry. d Representative images showing GFP positive HeLa-TL cells 24, 48 and 72 h after exposure to VFIC EVs at different doses. Scale bar, 100 µm. e–h Recombination in hard-to-transfect reporter cells (MSC-TL, THP-1-TL, Raw264.7-TL and K562-TL) mediated by VFIC EVs after 48 h. i The efficiency of Cre transfer in reporter cells by EVs isolated using TFF + UF + SEC method. TFF: tangential flow filtration; UF: ultrafiltration; SEC: size exclusion chromatography. j Dynamic Cre delivery in HeLa-TL cells using VFIC EVs purified by DGUC. DGUC: density gradient ultracentrifugation; T1-T3: technical replicates. Two-way ANOVA (Tukey) multiple comparisons test was used for analysis of (c), (e–h) and (i). Experiments were done with 3 biological replicates and data are shown as mean ± SD. Exact statistical analysis was reported in the Source data and Source data are provided as a Source Data file.

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