Optimized Protocol for the Isolation of Extracellular Vesicles from the Parasitic Worm Schistosoma mansoni with Improved Purity, Concentration, and Yield

Figure 1

Adult worm EV isolates prepared by differential (ultra)centrifugation (dUC) or size exclusion chromatography (SEC) are contaminated by hemozoin. Culture supernatant of adult worms was subjected to dUC (a, b, and e) or SEC (c, d). (a) Western blot detection of the schistosome EV marker tetraspanin-2 (TSP-2) in 100,000 × g pelleted adult worm EVs. (b) Black 100,000 × g pellet of adult worm EVs indicates the presence of hemozoin. Collected SEC fractions from adult worm E/S released by ~100 worms show similar fractions containing TSP-2 (c) and hemozoin (brown fractions in d). (e) Cryo-EM analysis of washed 100,000 × g pellet from 22 ml adult worm culture shows EVs (white arrowheads) and coisolated hemozoin crystals (black arrows). (f) Cryo-EM analysis shows that pelleted schistosomula EVs from 15 ml culture medium, with characteristic “hair-like” filaments on their surface, do not contain hemozoin crystals. Scale bars (e, f) are 500 nm.

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