pstat1 py701 antibody  (Cell Signaling Technology Inc)

Journal: Cell Reports Medicine

Article Title: Lymph node and tumor-associated PD-L1 + macrophages antagonize dendritic cell vaccines by suppressing CD8 + T cells

doi: 10.1016/j.xcrm.2023.101377

Figure Lengend Snippet:

Article Snippet: PE anti-mouse pSTAT1 (clone: pY701) , BD Biosciences , Cat#562069 RRID: AB_11151907.

Techniques: Recombinant, Lysis, Protease Inhibitor, Western Blot, Staining, Stripping, Liposomes, CRISPR, MTS Assay, ATP Assay, Cell Isolation, Enzyme-linked Immunosorbent Assay, Conjugation Assay, Selection, Vaccines, Single-cell Analysis, RNA Sequencing Assay, Mutagenesis, Microarray, Purification, Software

Journal: PLoS ONE

Article Title: Activation Mechanisms of Natural Killer Cells during Influenza Virus Infection

doi: 10.1371/journal.pone.0051858

Figure Lengend Snippet: NK cells from infected (open histograms) and uninfected (shaded histograms) mice were analyzed for intracellular pSTAT1 following adoptive transfer (A) or co-culture (B). Adoptive transfer was as described in . For in vitro infection, CD45.2 + splenocytes from IFNAR +/− or IFNAR −/− mice were combined with CD45.1 + B6 splenocytes at 1∶1 ratio, then infected with flu. NK cells (NK1.1 + CD3 − ) from infected (open histograms) and uninfected (shaded histograms) samples were analyzed for intracellular pSTAT1. Values represent the percentages of pSTAT1 + NK cells. Data are representative of three independent experiments with 2–4 (A) or 1–3 (B) mice per group.

Article Snippet: Antibodies against the following proteins were purchased as indicated: NK1.1 (PK136), CD3 (145-2C11), CD19 (1D3), IFN-γ (XMG1.2), granzyme B (GB11), CD107a (1D4B), CD69 (H1.2F3), pSTAT1 pY701 (4a) and pSTAT4 pY693 (38/p-Stat4) from BD Biosciences, CD49b (DX5) from eBiosciences, and CD45.2 (104) from Biolegend.

Techniques: Infection, Adoptive Transfer Assay, Co-Culture Assay, In Vitro

Journal: Frontiers in Immunology

Article Title: Type I interferon subtypes differentially activate the anti-leukaemic function of natural killer cells

doi: 10.3389/fimmu.2022.1050718

Figure Lengend Snippet: IFNβ, IFNα6 and IFNα14 activation induces stronger phosphorylation of STAT1 and STAT3 in NK cells than the clinical standard IFNα2B PBMCs were assessed for the expression of phosphorylated STAT1 (pY701) or STAT3 (pY705) at baseline (unstimulated) and following stimulation with 100 IU mL -1 of IFNβ, IFNα2B, IFNα6 or IFNα14 for 30 mins. Representative histogram plots show per-cell pSTAT1 or pSTAT3 expression in CD56 + CD3 - NK cells. Summary data show mean ± SEM pSTAT1 or pSTAT3 MFI in unstimulated and IFN-I stimulated NK cells. n = 7 healthy donors, 2 independent experiments. Data were compared using a paired Student’s t test. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001.

Article Snippet: The following antibodies were used: anti-CD56 (B159), anti-CD3 (SK7), anti-pSTAT1 (pY701), anti-pSTAT3 (pY705) and anti-pSTAT5 (pY694) (all BD Biosciences).

Techniques: Activation Assay, Expressing

Journal: Cell Reports

Article Title: Structural basis of activation and antagonism of receptor signaling mediated by interleukin-27

doi: 10.1016/j.celrep.2022.111490

Figure Lengend Snippet: High-affinity binding of gp130 to site 3 occurs independently of site 2 (A) Predicted location of the gp130 receptor binding consistent with the site 3 mode of interaction. Top panel: crystal structure of the mIL-27:mIL-27Rα:Nb5 complex (Nb5 not shown), bottom panel: crystal structure of the hIL-27:SRF388 Fab complex (SRF388 Fab not shown). (B–E) (B) Kinetic profiles of the receptor complex mediated by mIL-27 characterized by BLI. Biotinylated mIL-27 sc comprising the EBI3 subunit fused to the p28 subunit via a (GGGS) 4 linker (see ), was coupled to the surface of streptavidin-coated BLI sensors, followed by binding measurements in different concentrations of mIL-27Rα CHR , (C) mgp130 IgCHR in the presence of 100 nM of mIL-27Rα CHR , (D) mgp130 IgCHR , and (E) mgp130 CHR . Data traces (black) were fitted using a 1:1 interaction model (red) to quantify the kinetics ( k a , k d ) and binding affinity ( K D ) of the interactions using the Octet Analysis Studio 12.2.1.24 software. For each experiment three technical replicates were performed. The reported K D , k a , and k d values represent average values from three technical replicate experiments. (F) The anti-gp130 Ig antibody B-T2 blocks IL-27 signaling in U937 cells and (G) PBMCs. U937 cells or Ficoll-isolated human PBMCs were cultured in RPMI with various concentrations of anti-gp130 antibodies and rhIL-27 for 20 min at 37°C. Cells were fixed and stained for pSTAT1 (pY701). Samples were washed with FACS buffer, read on an LSR Fortessa (BD Biosciences), and analyzed using the FlowJo Software analysis program (TreeStar). Cytokine stimulated conditions represents 0% inhibition and unstimulated conditions represents 100% inhibition. The data presented represent the mean of two technical replicates per data point with error bars indicating SD. These data are representative of two independent experiments with U937 cell line or PBMCs from healthy donors.

Article Snippet: pSTAT1 (pY701) PE , BD Biosciences , Cat# 562069; RRID: AB_11151907.

Techniques: Binding Assay, Software, Isolation, Cell Culture, Staining, Inhibition

Journal: Cell Reports

Article Title: Structural basis of activation and antagonism of receptor signaling mediated by interleukin-27

doi: 10.1016/j.celrep.2022.111490

Figure Lengend Snippet:

Article Snippet: pSTAT1 (pY701) PE , BD Biosciences , Cat# 562069; RRID: AB_11151907.

Techniques: Recombinant, Plasmid Preparation, Inhibition, Software