Fig. 1: Isolation and characterization of ASCs and ASC-derived exosomes (ASC-Exos) from deer antler tissue.

A Schematic illustration of the ASCs isolation procedure from deer antler tissue using mechanical dissection and the collagenase digestion method. Created in BioRender. Yuhao, S. (2025) https://BioRender.com/m5easfx. B Representative phase-contrast microscopy image showing the typical spindle-shaped morphology of cultured ASCs at passage 3. Scale bar: 100 μm. C Immunofluorescence analysis of ASC surface markers. ASCs were positive for CD73 and CD90 (green), while negative for CD34 and CD45. Nuclei were counterstained with DAPI (blue). Merged images show co-localization. Scale bar: 100 μm. D–F Multi-lineage differentiation potential of ASCs. D Oil Red O staining showing lipid droplets after 14 days of adipogenic induction. Scale bars: 100 μm. E Alizarin Red S staining demonstrating calcium deposition after 21 days of osteogenic differentiation. Scale bars: 500 μm. F Alcian Blue staining revealing proteoglycan synthesis after 14 days of chondrogenic induction. Scale bars: 100 μm. G Western blot analysis confirming the expression of exosomal markers (CD63, TSG101, and CD9) in isolated ASC-Exos compared to ASCs and culture supernatant. H Representative transmission electron microscopy (TEM) image showing the typical cup-shaped morphology of ASC-Exos with characteristic double-membrane structure. Scale bars: 100 nm. I Nanoparticle tracking analysis (NTA) reveals the size distribution of ASC-Exos with an average diameter of 86.4 nm. J Scatter plot from flow cytometry analysis demonstrating the uniform distribution and homogeneity of the isolated ASC-Exos population.