Fei tecnai g2 twin

The isolated products from each experiment were processed in the same way. Paraformaldehyde was first added to the sample with a final concentration of 4% (w/v) and then incubated for 20 min at room temperature. A 100-μl droplet of isolated exosome sample was then placed on a sheet of Parafilm (VWR, USA). A 300-mesh copper grid support film (Electron Microscopy Sciences, USA) was placed on the droplet with membrane side down to allow the sample absorption for 20 min. Then, this grid was transferred to a 100-μl droplet of distilled water for 2 min. This process was repeated three times. This grid was then transferred to a 100-μl droplet of uranyl-acetate solution for negative staining for 8 min. Last, the grid was rewashed using distilled water and left to dry at room temperature. The sample was finally observed under TEM (FEI Tecnai G2 Twin, FEI Company, USA).

Microscopic images and videos were acquired using an upright BX51WI microscope (Olympus, Japan) combined with a CoolSNAP HQ2CCD camera (Photometrics, USA). The high-speed videos needed to capture the droplet rotation speed (Fig. 2), and an inverted microscope (TE2000-U, Nikon, Japan) equipped with a fast camera (Photron, Japan) was used. The droplet spinning motion was captured with a frame rate of 3000 fps and analyzed using ImageJ (National Institutes of Health (NIH), USA) and MATLAB R2016b (MathWorks, USA). The sEV samples were collected and visualized using TEM (FEI Tecnai G2 Twin, FEI Company, USA) with a negative staining method. The nanoparticle size distribution and concentration pre- and postprocessing were analyzed using a Malvern Zetasizer (Malvern Instruments, UK) and NTA with a NanoSight LM10 apparatus (Amesbury, UK) following the manufacturers’ protocols. For Zetasizer analysis, the samples were diluted 40× prior to analysis. For NanoSight analysis, 300 µL of the undiluted samples were directly loaded into the sample chamber using a 1 mL syringe. The camera level was set to 16, and the slider shutter was set to 1300. The NanoSight LM10 recorded 10- and 60-second videos for each sample, which were then analyzed using nanoparticle tracking analysis (NTA) 2.0 Analytical software. For sample analysis, the detection threshold was set to level 3.