mab3050  (R&D Systems)

Journal: Nature immunology

Article Title: Fungal sensing by dectin-1 directs the non-pathogenic polarization of T H 17 cells through balanced type I IFN responses in human DCs.

doi: 10.1038/s41590-022-01348-2

Figure Lengend Snippet: Fig. 6 | Dectin-1 induces αvβ8 expression via IFN-β, which is critical for TGF-β activation and non-pathogenic TH17 polarization. a,b, SEAP assay on supernatant of HEK-Blue TGF-β reporter cells for quantification of active TGF-β in the supernatant of unstimulated DCs or after stimulation of DCs with curdlan or C. albicans, in the presence of blocking αvβ1, αvβ3, αvβ5, αvβ6 and αvβ8 antibodies or isotype (IgG1 and IgG2a) control antibodies (a) or after transduction with non-targeting (control) or specific siRNAs to silence αv (ITGAV) or β8 (ITGB8) expression (b) at 24 h (a, IgG1, αvβ1, αvβ3, αvβ5, αvβ6 Ab n = 2, IgG2a, αvβ8 Ab n = 5; b, n = 4). Data in a,b represent mean ± s.d. of independent donors. **P < 0.01, *P < 0.05 (paired, two-tailed Student’s t-test), calculated between untreated and treated (a) or control and specific siRNA-transduced (b) samples that were likewise stimulated. c,d,g,h, Flow cytometry analyses by staining for αv (c) or β8 (d,g,h) expression (FI) in unstimulated DCs or after stimulation of DCs with curdlan or C. albicans (d), in the presence of blocking dectin-1 or IFN-α/ βR antibodies (g) or after silencing of IRF1 or IRF5 expression (h), at indicated times (c, n = 2; d,g, n = 3; h, n = 4). Isotype indicates negative control staining. Representative histograms for independent donors are shown. e,f, Real-time PCR analyses of ITGB8 relative mRNA levels in unstimulated DCs or after stimulation of

Article Snippet: DCs were preincubated for 2 h with MMP14 inhibitor NSC405020 (100 μM; Tocris) or blocking antibodies, anti-dectin-1 (20 μg ml−1; clone 259931, MAB1859, R&D Systems), anti-IFN-α/βR2 (20 μg ml−1; clone MMHAR-2, PBL Assay Science), anti-αvβ1 (10 μg ml−1; clone P5D2, MAB17781, R&D Systems), anti-αvβ3 (10 μg ml−1; clone 23C6, MAB3050, R&D Systems), anti-αvβ5 (10 μg ml−1; clone P5H9, MAB2528, R&D Systems), anti-αvβ6 (10 μg ml−1; clone 10D5, ab77906, Abcam), anti-αvβ8 (10 μg ml−1; kind gift from S.L.

Techniques: Expressing, Activation Assay, SEAP Assay, Blocking Assay, Control, Transduction, Two Tailed Test, Flow Cytometry, Staining, Negative Control, Real-time Polymerase Chain Reaction