Fig. 3.

Alum-induced PD1 limits CTL differentiation. (A) B6 mice were primed with OVA plus alum, MPL, or both adjuvants i.p. On day 7, these mice and naïve mice were injected with a 1:1 mixture of splenocytes, half of which had been incubated with SIINFEKL peptide and stained with a high level of CFSE and half of which had been stained with a low level of CFSE. Two days later, the percentage of total CFSE⁺ splenocytes that were CFSE^hi was examined. A representative sample from each group is shown. The number indicates the percentage of CFSE^hi cells that had been labeled with SIINFEKL out of total CFSE⁺ cells. The data are representative samples from two experiments with three mice per group. (B) B6 mice were primed with OVA plus alum, MPL, or both adjuvants i.p., and the levels of PD1 expression on the surface of K^b/SIINFEKL tet⁺ cells were examined on day 8. Cells were gated on K^b/SIINFEKL tet⁺ cells from the indicated mice (open histograms) or on naive CD8 T cells (filled histogram). (C) The MFI of PD1 on K^b/SIINFEKL tet⁺ cells was determined after immunization. *P < 0.05; **P < 0.01; ***P < 0.001. The data were combined from between one and three experiments with three or four mice per group for each time point. (D) B6 mice immunized with OVA plus alum were given anti-PDL1 or an isotype control antibody on days 0, 3, and 7 and SIINFEKL pulsed and unpulsed CFSE-labeled cells on day 7. Two days later, the percentage of CFSE^hi/ SIINFEKL pulsed cells out of the total CFSE⁺ cells was examined. Each point represents a mouse, and the line indicates the mean value for the group, combined from two experiments.