FIGURE 2.

The promising universal method applied for extracting Ti-EVs from a variety of tissues. At first, fresh tissues are sliced into slices and subsequently dissociated gently for 15–45 min at 37°C. Afterwards, a 70 μm pore-sized filter is used to remove tissue pieces. Centrifugation at a speed of 300–10,000 g at 4°C is conducted to remove apoptotic bodies, remaining cells and debris and the supernatant pass through a 0.22 μm filter to collect large EVs. Next, the filtrate is ultracentrifugated at a speed of 100,000–150,000 g at 4°C (>1 h) to collect small EVs. Finally, the isolated Ti-EVs are resuspended with PBS and can be stored at −80°C for a long time.