The retinas from 4–6-week-old wild-type C57BL/6J mice were extracted and digested using the Papain Dissociation System (Worthington Biochemical Corporation, Lakewood, NJ, USA) according to the manufacturer’s protocol. Exosomes were recovered using differential ultracentrifugation as previously described [26 (link)]. In brief, the supernatant received from the digestion process was mixed with cold HBSS and centrifuged at 25,000× RPM for 60 min at 4 °C in a Beckman Coulter Optima L80-XP ultracentrifuge with rotor SW60 Ti. The supernatant was again subjected to sequential ultra-centrifugation at 41,000× RPM for 60 min at 4 °C for 120 min. The final pellet (exosomes) was resuspended in 100 μL PBS and stored at −80 °C until further use (Figure 1 A). The size, number, and morphology of exosomes were characterized by nanoparticle tracking analysis (NTA) (NanoSight, Malvern Instruments Ltd., Malvern, UK) and transmission electron microscopy (TEM), respectively (Figure 1 B,D). Exosome markers were analyzed by an exosome detection antibody array (Exo-Check, System Biosciences, Palo Alto, CA, USA) according to the manufacturer’s instructions (Figure 1 C). Zeta potential of exosomes was measured using ZetaPALS (Brookhaven Instruments, Holtsville, NY, USA) (Figure 1 E).
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