Figure 2. Negative immune checkpoint blockade combined with mIL-15 down-regulated surface PD-1 expression on CD8⁺ T-cells and CD8⁺CD44^high T-cells that had been mediated by mIL-15.

Flow cytometry was used to analyze the PD-1 expression on splenic CD8⁺ T-cells as well as on the CD8⁺CD44^high population from the CT26-bearing mice following treatment with PBS, mIL-15, mIL-15 and anti-PD-L1, or mIL-15, anti-PD-L1 and anti-CTLA-4 on day 10 after CT26 tumor inoculation. Surface expression of PD-1 on CD8⁺ T-cells was detected using APC-anti-CD8 and PE-anti-PD-1 (open area, Fig. 2A). The CD8⁺CD44^high population was stained with APC-anti-CD8 and PE-Cy5.5-anti-CD44 antibodies (open area, Fig. 2B). An isotype matched IgG was used as a negative control (filled area). All the samples were analyzed on gated CD8⁺ T-cells or CD8⁺CD44^high cells as indicated. MFI is shown. The figure is representative of two independent experiments wherein each group contained three to five animals. Statistical analysis was performed based on PD-1 expression on CD8⁺ T cells (Fig. 2C) and CD8⁺ CD44^high population (Fig. 2D). * P<0.05 and ** P<0.01.