Research Article

Proteolytic Potential of the MSC Exosome Proteome: Implications for an Exosome-Mediated Delivery of Therapeutic Proteasome

Figure 4

Human plasma contained exosome-associated 20S proteasome. (a) Human plasma was loaded on a sucrose density gradient prepared by layering 14 sucrose solutions of concentrations from 22.8 to 60% (w/v) in a SW60Ti centrifuge tube and then ultracentrifuged for 16.5 h at 200 000 g, 4°C, in a SW60Ti rotor. 330 μL fractions were removed sequentially from the top and the density of each fraction was calculated by weighing a fixed volume of each fraction. The fractions were analyzed by Western blot analysis for PSMA1–7. (b) Relative amount of CD81 and PSMA1–7 in plasma exosome. Exosomes which are known to have membrane enriched in GM1 gangliosides were isolated from the sucrose density gradient by the high affinity of GM1 gangliosides for cholera toxin B (CTB) chains conjugated to magnetic beads. The CTB extract was assayed by ELISA for the amount of PSMA1–7 and CD81. Each point represents mean ± SEM ( ).