Anti migm

Pan B cells were isolated from spleens of Sox8^−/− or littermate control mice using MojoSort mouse pan B cell isolation kit (BioLegend) in accordance with the manufacturer’s instructions. For IgA⁺ B cell polarization, pan B cells were stimulated with 2 µg/ml F(ab′)2-goat-anti-mIgM (Thermo Fisher Scientific), 5 µg/ml anti-mCD40 (3/23; BioLegend) in complete advanced RPMI 1640 media (Thermo Fisher Scientific) containing 5% vol:vol FCS (MP Biomedicals) supplemented with 1 ng/ml recombinant human (rh) TGF-β1, 5 ng/ml recombinant mouse (rm) IL-5, 20 ng/ml rmIL-21 (all from BioLegend), and 10 nM all-trans retinoic acid (Fujifilm Wako Pure Chemical) for 6 d. For IgG1⁺ and IgE⁺ B cell polarization, pan B cells were stimulated with anti-mIgM and anti-mCD40 in complete media supplemented with 20 ng/ml rmIL-21 and 50 ng/ml rmIL-4 (BioLegend) for 6 d. For IgG2a⁺ B cell polarization, pan B cells were stimulated with anti-mIgM and anti-mCD40 in complete media supplemented with 20 ng/ml rmIL-21 and 50 ng/ml rmIFN-γ (BioLegend) for 6 d.