Fig 1: Bead-based analysis of phosphatidylserine (PS)-exposing sEVs and m/lEVs. (A) A schematic diagram of EV capturing on beads and analysis of their PS exposure by fluorescein isothiocyanate (FITC)-GlaS and allophycocyanin (APC)-annexin A5 (ANX5) proteins. (B–E) The beads capturing small EVs (sEVs) or medium/large EVs (m/lEVs) were labeled with FITC-GlaS or APC-ANX5 and analyzed by flow cytometry. The graphs show % of the FITC or APC positive EV-coated beads. Error bars, the standard deviation of five independent analyses, ** p < 0.01; *** p < 0.001; **** p < 0.0001.
Fig 2: Single EV analysis of phosphatidylserine (PS)-exposing small EVs (sEVs) and medium/large EVs (m/lEVs). (A) A schematic diagram of EV labeling with CellTrace Violet (CTV), followed by incubating with antibodies against EV markers and PS-binding proteins. (B,C) CTV-sEVs were incubated with allophycocyanin (APC)-anti-CD63 antibodies and fluorescein isothiocyanate (FITC)-GlaS or FITC-annexin A5 (ANX5) proteins. (D,E) CTV-m/lEVs were incubated with APC-anti-annexin A1 (ANX1) antibodies and FITC-GlaS or FITC-ANX5 proteins. The graphs show % of the FITC/APC double-positive EV populations. Error bars, the standard deviation of three independent analyses, * p < 0.05; ** p < 0.01; *** p < 0.001.
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