Transmission electron microscopy (TEM; Phillips CM120) was used to detect M2 BV2 cell-derived sEVs. sEVs were fixed in 2% paraformaldehyde on formvar-carbon-coated grids, and the membranes were covered for 30 min in a dry environment. Thereafter, the samples were washed with 100 μL of PBS, and grids were transferred to 50 μL 1% glutaraldehyde for 10 min before being transferred to 100 µL distilled water for 10 min. These steps were repeated seven times, after which the grids were dried and observed by TEM at 200 kV.

NTA was used to track the sEV diameter and number of particles. To provide representative size distribution profiles, data obtained from NTA were averaged within each sample across the video replicates and then averaged across samples. These distribution profiles were normalized to the total nanoparticle concentrations.

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