fig1

Figure 1. MEG-01 cells lack uniform expression of canonical EV tetraspanins, while maintaining platelet specific markers. (A) Representative bright-field microscopy image of MEG-01 cells culturing in serum-free media. Scale bar 100 μm; (B) Quantification of cell concentration at day 0 and day 2 for cells cultured in either full media (blue) or serum-free media (pink). Statistical significance was determined using an paired two-tailed Student’s t-test; P < 0.05. A significant increase in cell concentration was observed in full media, but not in serum-free media; (C) Flow cytometry dot plots showing Annexin V-FITC and PI staining of cells cultured for 2 days in full media or serum-free media; (D) Quantification of viable cells (Annexin V^-/PI^-; left) and apoptotic/necrotic cells (Annexin V⁺/PI⁺; right) from panel C, showing no significant difference between conditions; (E) Flow cytometry histograms showing the expression of surface markers CD9, CD63, CD81, CD41/CD61, CD40, CD41, CD42b, and CD62p on cells cultured in serum-free media. Blue histograms represent antibody staining and red histograms represent isotype controls. Representative bright-field images (A), flow cytometry plots (C), and flow cytometry histograms (E) from one of three independent biological experiments with similar results are shown. Quantitative data in (B and D) are presented as mean ± SD from three independent biological experiments. Statistical significance was determined using an unpaired two-tailed Student’s t-test. ns, not significant; ^***P < 0.001. APC: Allophycocyanin; EV: extracellular vesicle; FITC: fluorescein isothiocyanate; PI: propidium iodide; PerCP-Cy5.5: peridinin-chlorophyll-protein complex-cyanine 5.5; SD: standard deviation; AF750: Alexa Fluor 750.