Cd45rb

For surface staining, cells were stained with antibody to the following markers: CD45.2, CD4, CD19, NK1.1 CD11b, F4/80, CD44, and CD45RB (BioLegend, San Diego, CA). For intracellular cytokine staining, immediately after isolation, cells were cultured in RPMI complete medium and stimulated with phorbol myristate acetate (50 ng/mL) and ionomycin (1 μg/mL) in the presence of brefeldin A (10 μg/mL; eBioscience, San Diego, CA) for 4 hours. Cells were washed and subjected to surface staining. Cells were then fixed and permeabilized using eBioscience kit following manufacturer’s guidelines. Mouse phycoerythrin-IL22 (eBioscience), allophycocyanin-IL17α (BioLegend), AF700-interferon gamma (BD, San Jose, CA), and matched isotype controls were used for intracellular staining. For human T-cell intracellular staining, phycoerythrin-IL22, allophycocyanin-IL17α, and AF700-interferon gamma antibodies were all from BioLegend. Dead cells were excluded from analysis using violet viability stain (Invitrogen). Flow cytometry data collection was performed on Fortessa LSRII (BD) and analyzed using FlowJo software (Tree Star, Inc, Ashland, OR).

Isolation of pancreatic leukocytes was through collagenase digestion of mouse pancreas as previously described^{13 (link)}. Dead cells were stained with LIVE/DEAD™ Fixable Aqua Dead Cell Stain Kit (Thermofisher scientific, Santa Clara, CA). For surface staining, cells were stained with antibody to the following markers: CD45, CD4, CD8, CD11b, F4/80, CD11c, CD44 and CD45RB (BioLegend, San Diego, CA). For intracellular staining, cells were stained with IFNγ, IL-4 and foxp3 (Biolegend, San Diego, CA), IL-17A and RORγt (eBioscience, San Diego, CA). Intracellular STING staining was as previously described¹⁰ , cells were stained with surface markers first, then fixed and permeabilized with a kit reagents from eBioscience (San Diego, CA). Then Rabbit unconjugated STING antibody or Rabbit IgG isotype control (Invitrogen, Carlsbad, CA) were used as primary antibody and AF488 conjugated goat anti-rabbit was used as secondary antibody (Life Technologies, Carlsbad, CA).