Immune cells were stained using antibodies against FITC-conjugated anti-mouse CD19 (ID3, TONBO Bioscience, San Diego, CA, USA), CD44 (IM7, eBioscience, San Diego, CA, USA), and Foxp3 (FJK-16s, eBioscience), PE-conjugated anti-mouse CD8 (53-6.7, eBioscience), PE-Cy7-conjugated anti-mouse CD4 (GK1.5, TONBO Bioscience), APC-conjugated anti-mouse CD62L (MEL-14, Invitrogen, Carlsbad, CA, USA), and APC-Cy7-conjugated anti-mouse CD45.2 (A20, BioLegend). A FACSCanto flow cytometer (BD Biosciences, San Jose, CA, USA) was used to quantify cell populations based on expression profile. Isotype antibodies were used for analysis of each surface molecule as a control. Immune cells obtained from spleen and salivary gland tissues in NOD mice were analyzed according to a gating strategy as previously described [38 (link)]. Briefly, lymphocyte subset was checked by gating on side scatter (SSC)/forward scatter (FSC). Then, single cells were obtained by discriminating doublets using FSC-Hight (H)/FSC-Area (A), and viable immune cells were gated on CD45.1+ and 7-aminoactinomycin D (7AAD)− (Invitrogen). Data were analyzed using the FlowJo FACS Analysis software (BD Biosciences).
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