Confirmation of successful plasma exosome enrichment from male Fisher 344 rats.

<p>A. TEM demonstrates successful isolation of exosome-sized particles (∼80 nm) from the plasma with ExoQuick precipitation. B. TEM shows how pre-treating plasma with Thromboplastin-D prior to exosome isolation with ExoQuick removes clouding factors and prevents vesicle aggregation. C. Exosome enriched and exosome depleted plasma fractions from stressed rats pre-treated with Thromboplastin-D were characterized for size distribution and quantitated by NanoSight LM10, using light scatter from a 532 nm green laser. ELISA analyses reveal that ExoQuick exosome isolation successfully enriches exosomes as marked by known exosome markers. Exposing rats to tail shock stress (Stress) had no impact on the rate of plasma exosome release compared to non-stressed rats (Control) as measured by: D. the tetraspanin CD63, E. the adhesion molecule Rab5b, F. and the intestinal exosome marker A33. G. Analysis of IL-6 confirms activation of the stress response and specificity of ExoQuick for exosome associated proteins. These data suggest that stress does not impact the rate of exosome release into the plasma. Proteomic results are expressed in means ± SE; 6–8 rats/condition. *indicates significant difference when compared to control rats (p<0.05). #indicates significant difference when compared to exosome depleted fraction (p<0.05). Two-way ANOVA with Fisher PLSD post hoc test was used.