Clone tc11 18h10

Mice were administrated 50 μg/mouse of an IL-7-neutralizing antibody (BioXCell, clone M25) at days 27, 30, 32, and 34 of the first immunization (day 0) [30 (link)]. IL-17A was blocked using 300 μg anti-mouse IL-17A mAb [31 (link)] (BioLegend, Clone TC11-18H10.1) administered i.v. into mice 2 d before P. aeruginosa XN-1 infection (at days 33 and 34). For neutralization of IFN-γ, mice were given intravenous injection 2 days of 300 μg anti-mouse IFN-γmAb [32 (link)] (BioLegend, Clone R4-6A2) before P. aeruginosa XN-1 infection (at days 33 and 34).

CD4⁺ T cells-depleted splenic cells were cultured with IL-23 (20 ng/ml, Biolegend) with or without butyrate (0.5 mM) for 16 h, and then stimulated with ionomycin (750 ng/ml) and phorbol-12-myristate 13-acetate (50 ng/ml) for 2 h, followed by addition of brefeldin (BD Biosciences) for another 3 h. After Fc blocking, cells were stained with surface markers (PE/Cy7-anti-Thy1, 1:200, Clone#30-H12, Cat#105326; FITC-lineage (CD3, Clone#145-2C11, Cat#100306; CD11b, Clone#M1/70, Cat#101206; CD11c, Clone#N418, Cat#117306; B220, Clone#RA3-6B2, Cat#103206; F4/80, Clone#BM8, Cat#123108; NK1.1, Clone#PK136, Cat#108706; and Gr1, Clone#RB6-8C5, Cat#108419), 1:100), which were purchased from Biolegend. Cells were permeabilized using Foxp3/Transcription Factor Fixation/Permeabilization set (Cat#00-5523-00, ThermoFisher), followed by intracellular staining (anti-IL-22, 1:200, Clone#1H8PWSR, Cat#12-7221-82, Invitrogen; anti-IL-17, 1:200, Clone#TC11-18H10.1, Cat#506916, Biolegend). For ILC3, cells were also stained with RORγt (1:200, Clone#B2D, Cat#17-6981-82, Invitrogen).
All the events were collected by BD FACS Diva software. IL-22 and IL-17 production in ILCs (Thy1⁺ Lineage⁻ cells), or in ILC3 (Thy1⁺ Lineage⁻ RORγt⁺ cells) was analyzed using FlowJo. Gating strategies are included in Supplementary Fig. 15.

Non-parenchymal liver cells were incubated with monoclonal antibody 2·4G2 for FcR blocking (BioLegend, San Diego, CA, USA) and then exposed at 4°C to a mixture of the following antibodies (dilutions are indicated):anti-mouse CD45 (clone 30-F11, 1:100), anti-mouse/human CD11b (clone M1/70, 1:300), anti-mouse Ly6C (clone HK1.4, 1:300), anti-mouse MHCII (clone M5/114.15.2, 1:200), anti-mouse CD11c (clone N418, 1:100), anti-mouse CD3ϵ (clone 145-2c11, 1:100), anti-mouse CD8a (clone 53-6.7, 1:100), anti-mouse CD4 (clone GK1.5, 1:100), anti-mouse TCRβ (clone 457-597, 1:100) all were purchased from BioLegend, San Diego, CA. Anti-mouse F4/80 (clone REA 126, 1:100) and anti-mouse Tim4 (clone REA999, 1:100) were purchased from Miltenyi Biotech.
For IL-17A staining, cells were first stained for surface markers, then the cells were fixed and permeabilized prior to intra-cellular staining with IL-17 (Clone TC11-18H10.1, 1:50, BioLegend). Cells were analyzed with BD FACS Canto™ II (BD Bioscience) or sorted with a FACSAria flow cytometer (BD Bioscience). Flow cytometry analysis was performed using FlowJo software (TreeStar, Ashland, OR).