human ifnar blocking antibody  (R&D Systems)

Journal: Nature

Article Title: Lysine Catabolism Reprograms Tumour Immunity through Histone Crotonylation

doi: 10.1038/s41586-023-06061-0

Figure Lengend Snippet: a, IB analysis of GSC23 after GCDH KD at the indicated time. b, ELISA quantification of IFNα and IFNβ in culture supernatants from GSCs with or without GCDH KD. nd, not detected. c, d, IB analysis of GSC23 treated with IFNβ (5 ng/ml, c) or culture supernatants from GSC23 with or without GCDH KD (d) for indicated time. IFNβ was added every 2 days for the duration of experiments. The supernatants with or without IFNAR blocking antibody were replaced every 2 days. e, IF staining for IFNα, Kcr, Kac, Kglu and GCDH in indicated sections from GSC23-derived intracranial tumours (n = 3 biologically independent mice). Scale bar, 20 μm. f, RT-qPCR analysis of human ISGs in GSC23-derived intracranial tumour tissues (n = 4 biologically independent mice). g, The gating strategy of GSCs in flow cytometric analysis. h, i, Flow cytometry plots (h) and quantification (i, n = 4 biologically independent mice) of SOX2+ CD133+ GSCs in CD147+ human tumour cells as indicated. Representative of two independent experiments in a, c and d. Data are presented from three independent experiments in b. In b, f and i, data are presented as mean ± SEM. One-way ANOVA followed by multiple comparisons with adjusted p values for f and i.

Article Snippet: To block GCDH loss induced activation of IFN signalling, GSCs were preincubated with 10 μg/ml of human IFNAR blocking antibody (cat: MAB4015; R&D Systems) for 2 h in 37 °C, followed with stimulation of the culture supernatants from control or GCDH KD GSCs.

Techniques: Enzyme-linked Immunosorbent Assay, Blocking Assay, Staining, Derivative Assay, Quantitative RT-PCR, Flow Cytometry

Journal: PLoS ONE

Article Title: Chikungunya virus superinfection exclusion is mediated by a block in viral replication and does not rely on non-structural protein 2

doi: 10.1371/journal.pone.0241592

Figure Lengend Snippet: ( A – C ) HFFs were treated with human IFNAR blocking antibody or isotype control at 5 μg/mL for 1 h, then were infected with CHIKV-GFP for 24 h at the indicated MOI, followed by CHIKV-mCherry at MOI 1 for 24 h. Blocking antibody treatment was maintained throughout the experiment. Representative flow cytometry plots (A) and quantification of infected cells (C) and are shown. Arrows highlight the difference in GFP infection between isotype control and blocking antibody treated samples. MX1 RNA levels were assessed by RT–qPCR (B). ( D , E ) WT (D) or Irf3 −/− Irf7 −/− MEF cells (E) were infected with CHIKV-GFP at the indicated MOI for 16 h, then with CHIKV-mCherry at MOI 5 (WT) or 3 ( Irf3 −/− Irf7 −/− ) for 8 h, and subsequently analyzed by flow cytometry. Bars indicate mean and SD of biological triplicates, and data are representative of at least two independent experiments. NS, not significant; *** p < 0.001 (one-way analysis of variance followed by Dunnett’s post-test).

Article Snippet: Human IFNAR blocking antibody (PBL Assay Science #21385–1) was used at a concentration of 5 μg/mL, ActD (Sigma-Aldrich #A1410) at 2 μg/μL.

Techniques: Blocking Assay, Control, Infection, Flow Cytometry, Quantitative RT-PCR

Journal: Cell reports

Article Title: Constitutive Interferon Attenuates RIPK1/3-Mediated Cytokine Translation

doi: 10.1016/j.celrep.2019.12.073

Figure Lengend Snippet: (A) TNF-α and CXCL-1 protein levels after indicated stimulations ± treatment with eIF4E inhibitor 4EGi-1 in B6, Ifnb −/− , and RIP1 Ki Ifnb −/− BMDMs. (B) p-eIF4E and total 4E-BP levels in B6, Ifnb −/− , and RIP1 Ki BMDMs stimulated as indicated ± 5 IU IFNβ priming overnight. (C) TNF-α and CXCL-1 protein levels after indicated stimulations ± treatment with mTORC1 inhibitor Torin 2 in B6, Ifnb −/− , and RIP1 Ki Ifnb −/− BMDMs. (D and E) Cell death as measured by propidium iodide incorporation over 6 h in B6, Ifnb −/− , and RIP1 Ki Ifnb −/− BMDMs stimulated with LZ ± treatment with 4EGi-1 (D) or Torin 2 (E). (F) TNF-α and CXCL-1 protein levels after indicated stimulations ± overnight treatment with IFNAR-blocking antibody (αIFNAR) in Akt1 −/− and wild-type (WT) littermate BMDMs. In all panels, BMDMs were stimulated with LPS, LZ, LZNs, Z, Ns, or LZN as indicated. ELISA and qPCR data are shown as ±SD from three independent experiments compared using two-way ANOVA: n.s. (p > 0.05), *p < 0.05, and ****p < 0.0001. Western blot and kinetic cell death experiments are representative of three or more independent experiments. Kinetic data are presented as the mean ± SD of triplicate wells. See also and .

Article Snippet: Blocking antibody to Human IFNAR (21385–1) was purchased from PBL Assay Science and used at 20ug/ml.

Techniques: Blocking Assay, Enzyme-linked Immunosorbent Assay, Western Blot

Journal: Cell reports

Article Title: Constitutive Interferon Attenuates RIPK1/3-Mediated Cytokine Translation

doi: 10.1016/j.celrep.2019.12.073

Figure Lengend Snippet: (A) Cell death as measured by propidium iodide incorporation over 8 h in human PBMC-derived macrophages stimulated as indicated ± overnight treatment with IFNAR-blocking antibody. (B and C) CXCL-1 protein (B) and mRNA (C) levels in human PBMC-derived macrophages stimulated as indicated ± overnight treatment with IFNAR-blocking antibody (D and E) CXCL-1 protein (D) and mRNA (E) levels in human PBMC-derived macrophages stimulated as indicated ± overnight treatment with IFNAR-blocking antibody and/or treatment with 4EGi-1 or Torin 2. In all panels, human PBMC-derived macrophages were stimulated with LPS, LZ, or LZNs as indicated. ELISA and qPCR data are shown as ±SD from three independent experiments compared using two-way ANOVA: n.s. (p > 0.05), **p < 0.01, ***p < 0.001, and ****p < 0.0001. Kinetic cell death experiments are representative of three or more independent experiments and presented as the mean ± SD of triplicate wells.

Article Snippet: Blocking antibody to Human IFNAR (21385–1) was purchased from PBL Assay Science and used at 20ug/ml.

Techniques: Derivative Assay, Blocking Assay, Enzyme-linked Immunosorbent Assay

Journal: Cell reports

Article Title: Constitutive Interferon Attenuates RIPK1/3-Mediated Cytokine Translation

doi: 10.1016/j.celrep.2019.12.073

Figure Lengend Snippet:

Article Snippet: Blocking antibody to Human IFNAR (21385–1) was purchased from PBL Assay Science and used at 20ug/ml.

Techniques: Purification, Recombinant, Enzyme-linked Immunosorbent Assay, RNA Sequencing, Derivative Assay