Fig 1.
Exosomes can be isolated with 1h ultracentrifugation method from blood plasma.
(A) Transmission electron microscopy images of exosome isolates from rat and human blood plasma. (B) CD63, TSG101 and albumin content of the rat and human exosomal isolates as evaluated with Western blot. (C) Size distribution of particles isolated from rat and human blood plasma analyzed with dynamic light scattering (averages of n = 3–4).
Fig 2.
Efficiency of ultracentrifugation (UC) methods under various conditions.
(A) Scheme of serial UC of the supernatant. Exosomal marker CD63 and TSG101 and albumin content of exosomal isolates after serial UC of the supernatant with 1h UC at 4°C (B; n = 3; p>0.05), with 10-fold dilution at 4°C (C, top), with smaller volume of loaded sample at 4°C (C, middle) and with 1h UC at 37°C (C, bottom) as evaluated with Western blot.
Fig 3.
The effect of various ultracentrifugation (UC) duration on the exosomal yield and purity.
(A) Transmission electron microscopy images of rat exosomes isolated with 1, 3, 6 or 14h UC period. (B) Size distribution of rat exosomes based on transmission electron microscopy image analysis (1h UC: n = 2,440; 3h UC: n = 353). (C) Size distribution of particles isolated with different UC duration evaluated with dynamic light scattering (averages of n = 3). (D) Protein concentration of exosome homogenates isolated with different UC duration as assessed with bicinchoninic acid assay (n = 3–7; *, #, &: p<0.05 vs. other three groups). (E) CD63 and albumin content of exosome isolates with different UC length (n = 3; *, #, &: p<0.05 vs. corresponding other three groups; §: p<0.05 vs. 1h).
Fig 4.
Efficiency and selectivity of size exclusion chromatography (SEC) on exosome isolation performed with various matrices.
(A) CD63, TSG101 and albumin content of different fractions collected during SEC on Sepharose 2B (top), Sepharose CL-4B (middle) and Sephacryl S-400 (bottom) columns with equal volumes (left column) or equal protein amounts of fractions (right column) loaded for Western blot. (B) Size distribution of particles isolated with various SEC matrices evaluated with dynamic light scattering. Sepharose 2B (top), Sepharose CL-4B (middle) and Sephacryl S-400 (bottom) columns.
Fig 5.
Size exclusion chromatography isolation of exosomes from rat blood plasma on a large-scale Sephacryl S-400 column.
(A) Particle concentration and protein content (absorbance at 280 nm) of eluted fractions. (B) Concentration and size distribution of particles in pooled fractions by nanotracking analysis. (C) CD63, TSG101 and albumin content of pooled