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. 2020 Dec 17;4(24):6274–6282. doi: 10.1182/bloodadvances.2020003012

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Figure 2. — Isolation of EVs from human milk. Human milk was fractionated by Sepharose 2B SEC, and all fractions were blotted for common EV markers tetraspanins CD63 (A) and CD9 (B). CD63 blot was reused for CD9 staining. Starting material is shown, as well as platelet lysate (positive control) and human brain lysate (negative control). EV-containing fractions 8 and 9 were pooled, and presence of CD9 and CD63 were determined on single EVs by flow cytometry (C) and on bulk EVs by surface plasmon resonance imaging (D). Pooled fractions 25 and 26 were used as negative and procedural controls in panels C and D, respectively. Immunoglobulin G1 (IgG1) was used to correct for binding of antibodies to Fc receptors exposed on EVs. ***P < .001. LALS, large angle light scatter; SPRi, surface plasmon resonance imaging.