pstat3-pe antibody ebioscience #501122408  (Thermo Fisher)

Journal: Scientific Reports

Article Title: Unveiling novel insights into human IL-6 − IL-6R interaction sites through 3D computer-guided docking and systematic site mutagenesis

doi: 10.1038/s41598-024-69429-w

Figure Lengend Snippet: Activation of phosphorylation of STAT3 in HepG2 cells by hIL-6 or its mutants. HepG2 cells were treated with 50 ng/mL hIL-6 or its mutants for 15 min, and cell lysates were analyzed to detect phosphorylation of STAT3. Untreated cells were used as the negative control (NC). ( A ) Representative, digital western blot images of expression of P-STAT3, STAT3 and β-Actin upon stimulation with hIL-6 WT or R167A, E171A and R178E mutants, respectively; ( B ) statistical analysis of hIL-6 mutants compared with hIL-6 WT in term of STAT3 phosphorylation levels in HepG2 cells, based on three independent experiments. Data is presented as mean ± SD, n = 3. **p < 0.01; and ***p < 0.001.

Article Snippet: The antibody, PE labeled anti-pSTAT3 (pY705) monoclonal antibody was purchased from BD (Cat. 612569), and horseradish peroxidase (HRP)-conjugated goat anti-human IgG antibody was obtained from Southern Biotech (Cat. 2049–5).

Techniques: Activation Assay, Negative Control, Western Blot, Expressing

Journal: Scientific Reports

Article Title: Unveiling novel insights into human IL-6 − IL-6R interaction sites through 3D computer-guided docking and systematic site mutagenesis

doi: 10.1038/s41598-024-69429-w

Figure Lengend Snippet: Activate of phosphorylation of STAT3 in Leukocytes by IL-6 mutants. Human leukocytes were preincubated with 50 ng/mL IL-6 mutants for 15 min, followed by permeabilized and stained. Untreated cells were used as the negative control (NC). Phosphorylation of STAT3 was assessed using flow cytometry with PE labeled anti-pSTAT3 (pY705) monoclonal antibody. ( A – C ) Gating strategy and representative histogram graph of P-STAT3 + cells.; ( D ) statistical analysis of percentage of P-STAT3 + cells upon stimulation with hIL-6 WT or R167A, E171A and R178E mutants, based on three independent experiments. The data is presented as mean ± SD, n = 3. *p < 0.05; **p < 0.01.

Article Snippet: The antibody, PE labeled anti-pSTAT3 (pY705) monoclonal antibody was purchased from BD (Cat. 612569), and horseradish peroxidase (HRP)-conjugated goat anti-human IgG antibody was obtained from Southern Biotech (Cat. 2049–5).

Techniques: Staining, Negative Control, Flow Cytometry, Labeling

Journal: bioRxiv

Article Title: LDL-Binding IL-10 Reduces Vascular Inflammation in Atherosclerotic Mice

doi: 10.1101/2024.03.04.582839

Figure Lengend Snippet: Fusing IL-10 to an LDL-binding antibody fragment enables lipoprotein binding while maintaining signaling and anti-inflammatory properties. a) Conceptual schematic of plaque-targeted IL-10. b) SDS PAGE gel showing various Fab-IL-10 constructs. N, non-reducing conditions. R, reducing conditions. c-e) Binding affinity of Fab-IL-10 to LDL measured using surface plasmon resonance. KD, dissociation constant. Dashed lines represented calculated fit. f) Activity of IL-10 and Fab-IL-10 by phosphorylation of STAT3 (pSTAT3), measured by flow cytometry of RAW 264.7 cells incubated with the indicated concentrations of IL-10 (n = 3). g) Log(EC50) calculated from fitted curves in (f), shown with 95% confidence intervals. h) TNFα secretion of LPS-stimulated RAW 264.7 cells incubated with WT IL-10 or Fab-IL-10 (n = 5). Experiments were performed twice with similar results. Data represent mean +/-standard deviation (f) or mean + standard deviation (h). Statistics performed by one-way ANOVA with Dunnett’s post-test compared to media.

Article Snippet: Cells were stained with PE-conjugated antibodies against pSTAT3 (pY705, clone 4/P-STAT3, BD Biosciences) at a 1:50 dilution for 1 hr at room temperature in the dark.

Techniques: Binding Assay, SDS Page, Construct, SPR Assay, Activity Assay, Flow Cytometry, Incubation, Standard Deviation

Journal: Journal for Immunotherapy of Cancer

Article Title: Breast cancer-induced immune suppression in the sentinel lymph node is effectively countered by CpG-B in conjunction with inhibition of the JAK2/STAT3 pathway

doi: 10.1136/jitc-2020-000761

Figure Lengend Snippet: Effect of CpG±AG490 on pSTAT3 levels in BrC SLN-derived cDC and pDC. Phosphorylated signal transducer and activator of transcription 3 (pSTAT3) levels (assessed by flow cytometric analysis) in the conventional DC population (phenotypically characterized by shared CD11c hi ) and the pDC subset measured in metastasis negative BrC SLN cell samples, cultured for 24 hours with medium, CpG or CpG+AG490. (A) Representative histogram plots of pSTAT3 fluorescence intensity (FI) levels in CD11c hi CDC (24 hours) from one of three individual BrC SLN cell samples are shown. Positive FI ranges according to fluorescence minus one control values are displayed with gray markers in each histogram. Both the mean fluorescence intensity (MFI) values and percentage pSTAT3 positive cells are displayed in boxes. (B) MFI of pSTAT3 and percentage pSTAT3 positive cells (both at 24 hours of culture) of cDCs (top bar graphs) and pDCs (bottom bar graphs) in three individual BrC SLN cell samples. Bars represent mean with SEM, with mean pSTAT3 levels (±SEM) of each culture condition and corresponding p values displayed above the error bars. BrC, breast cancer; cDCs, conventional DCs; DC, dendritic cell; pDCs, plasmacytoid DCs; SLN, sentinel lymph node; STAT3i, STAT3 inhibitor.

Article Snippet: STAT3 phosphorylation (pSTAT3) was assessed by intracellular staining with the PE-conjugated mAb against pSTAT3 (pY705, clone 4/pSTAT3; BD Phosflow), following the manufacturer’s instructions.

Techniques: Derivative Assay, Cell Culture, Fluorescence, Control