Tcr β bv421

Single cells were harvested using 45 μm filters (Biologix, Shanghai, China) from spleen and draining lymph nodes at the peak of psoriasis. At first, cells were washed three times with PBS. Surface staining of innate immune cells was performed with fluorescent-labeled antibodies: F4/80-PerCP-Cy 5.5, CD11c-PE, CD11b-APC, and GR-1-FITC. Staining of γδ T cells and their subsets was performed with CD3-APC, γδ TCR-PE, TCR-β-BV421, Vγ4-FITC, and Vγ5-BV510 (BD Biosciences, New Jersey, USA) antibodies for 30 min at RT. FACS was done using LSR II (BD Biosciences) and data were analyzed using Flow Jo version 7.0 (Tree Star, California, USA).

CD1a-PE, CD1b-PE, CD1c-FITC, B220-PerCPCy5.5, F4/80-APC, CD11c- BV421, CD8-BV510, CD4-PerCPCy5.5, IFN-γ-APC, IL-17-PE, Granzyme B-BV421, and TNF-α-FITC antibodies were purchased from BioLegend (San Diego, CA). TCR-β-BV421, anti-Vβ3 and 4-PE, and anti-Vβ5.1/5.2-APC were obtained from BD biosciences (San Jose, CA). For flow cytometry, single cell suspensions from organs were incubated with 2.4G2 FcR blocking antibody for 10 minutes and then stained in HBSS-2% FBS containing 50 μg/mL gentamicin. Samples were run on the FACS Canto II flow cytometer (BD Biosciences, San Jose, CA). Analysis of flow cytometry data was performed on FlowJo software (Tree Star, Inc). S12 Fig shows the gating strategies employed for the FACS plots of group 1 CD1-restricted T cells in the polyclonal setting in Fig 3 and group 1 CD1-restricted T cell line functional assays in Fig 5.