Evo ls 15 scanning

Isolated EVs intended for scanning electron microscopy (SEM) were fixed with glutaraldehyde at a final concentration of 2.5% in filtered PBS and stored at 4 °C until further preparation. After homogenization, 10–20 mL of each sample was placed onto a Thermanox coverslip (Thermo Fisher Scientific, Waltham, MA, USA) and allowed to settle for 90–120 min in a humid chamber to prevent drying. For dehydration, the samples were then placed into solutions with increasing acetone concentration (70–100%) and subsequently fully dried via critical point drying using CO₂ to avoid shrinkage effects and loss of structure from air-drying. The dehydrated samples were sputtered with gold and analyzed with an EVO LS 15 scanning electron microscope (Carl Zeiss AG, Oberkochen, Germany).

A EVO LS15 scanning electron microscope (Zeiss, Germany) with Q150RS ion sputtering coater (Quorum, UK) were used to examine the morphological alterations of samples. The samples of before and after fermentation and the sample residue from each stage of in vitro digestion were dried at 60 °C and crushed. The crushed samples were fixed on the SEM sample stage with conductive adhesive and were coated with gold on the sample surface using an ion sputter coater. Then, the samples were observed at an accelerating voltage of 10 kv (2000× magnification).