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Whole blood was collected from C57BL/6J male mice (twelve weeks), and the serum was carefully separated, as previously described. The serum was centrifuged at 10,000 × g for ten minutes to remove cellular debris and subsequently ultracentrifuged at 210,000 × g for thirty minutes. The pellet was washed and resuspended in PBS solution. To characterize EVs collected from whole blood, nanoparticle-tracking analysis (NTA) was performed by using NanoSight (Figure 3.8). The peak size of the EVs was 67 nm, which was comparable with that described in previous reports (Lobb et al. 2015; Urabe et al. 2017).

Figure 3.8 Evaluation of EVs using NanoSight. Serum was separated after blood collection, and microparticle analysis by NanoSight was performed.