Fig. 5

CD8⁺ T cells demonstrate the suppression of proliferation of responder T cells (Tresp) after incubating them with 4 pg μl ⁻¹ TDEs and responder T cells: CD8+ T cells down to 512:1. a. CD8⁺ T cells were purified from human buffy coat and then incubated for 6 h with serially diluted concentration of 30,000 pg μl⁻¹ of TDEs from Tu167. TDE concentrations were determined from BCA protein assay. After the CGM from the tumor cell lines had dead cells and debris removed, it was incubated overnight at 4 °C with ExoQuick-TC (System Biosciences) to purify exosomes. This produced an exosome fraction (pellet) and a non-exosome membrane vesicle fraction (supernatant). After incubation cells were washed and grown in complete RPMI for 7 days. On day 6, a portion of CD8⁺ T cells were removed and used for the above suppression assay at a 2:1, Responder T cells: CD8+ T cells. A BrdU incorporation ELISA suppression assay was done for 72 h with BrdU being added 22 h before the end of the experiment. b. The above was repeated with Tu167 TDEs but at 30 ng μl⁻¹. On day 6, a portion of CD8+ T cells were removed and used for the suppression assay at ratios between 2:1 to 512:1, Responder T cells: CD8+ T cells. All data represents the mean +/− SD of 3. To compare means, a one-way analysis of variance was used with a Bonferroni multiple comparison test. ***p < 0.001 for the indicated groups vs. responder T cells (Tresps) alone