Cd196 pe

To identify specific immune cell subsets in PBMCs following VitD₃ treatment, cells were stained with fluorescently-conjugated monoclonal antibodies; CD4-BUV737, CD45RO-APC, CD161-FITC, CD194-V450, CD196-PE, CD14-BV605, CD19-APC-H7, CD56-BV421, CD282-AF647 (TLR2), CD284-BV786 (TLR4; all from BD Bioscience; San Diego, CA, USA), and anti-TLR7-PE (Gibco Life Technologies, Carlsbad, USA), Zombie Aqua™ Fixable Viability Kit (BioLegend, San Diego, USA). Compensation bead particles were used to account for spectral overlap (BD Bioscience, San Diego, CA, USA) and analyzed using the BD LSRII flow cytometer. Unstained PBMCs and fluorescence minus one (FMO) were used as controls and a minimum of 100,000 events were analyzed per sample gated on live, single cell lymphocyte gate based on FSC and SSC, where the expression of the cell surface molecules was evaluated using FlowJo, LLC v10.4.2 software. Refer to Supplementary Figures 1–4 for gating strategies.

The whole blood samples of 30 patients with PTB and 30 controls were collected. Cell surface markers were then stained with antibodies against CD4 FITC (Cat 555346), CD196 PE (Cat 559562), CD183 ECD (Cat 551128) from BD (San Diego, USA), according to standard procedures. The gating strategy was CD4 + CD183 + CD196- for Th1, CD4 + CD183− CD196 for Th2, and CD4 + CD183-CD196 + for Th17^{26 (link)}. The cells were treated with red blood cell lysate (BD, California, USA) and washed with PBS twice. Flow cytometry of cells was performed with a BD LSRII flow cytometer and analyzed with FlowJo software (v7).