Western Blot Analysis of EV Fractions

WB analysis was performed as described previously [85 (link)]. Equal amounts of EV fractions (15 µL (~10 µg); F1–F8) or whole-cell lysate (10 µg; prepared using RIPA buffer: ThermoFisher Scientific, Waltham, MA, USA) were solubilized using 4× Laemmli buffer (Biorad, Feldkirchen, Germany) in reducing conditions at 95 °C for 10 min. Samples were then separated by SDS-PAGE under reducing conditions, and immunoblot analyses were performed using antibodies (Supplementary Table S8) followed by horseradish peroxidase-labeled anti-mouse or rabbit IgG (Jackson ImmunoResearch). Protein bands were visualized using the enhanced chemiluminescence Western blot detection reagent (GE Healthcare) and the fusion fx Imager/fusion software (Fusion FX7 Edge 18.05, Vilber Lourmat; https://www.vilber.com/fusion-fx/).