rabbit polyclonal anti pstat1  (Cell Signaling Technology Inc)

Journal: Gut Microbes

Article Title: Synergistic activity of Enterococcus Faeciu m-induced ferroptosis via expansion of IFN-γ + CD8 + T cell population in advanced hepatocellular carcinoma treated with sorafenib

doi: 10.1080/19490976.2024.2410474

Figure Lengend Snippet: Efm enhanced the ability of sorafenib to induce ferroptosis in hepatocellular carcinoma. (a) The relative protein expression of SLC7A11, STAT1 and pSTAT1 were assessed by western blotting. (b,c) the binding sites between STAT1 and SLC7A11 predicted by JASPAR database. (d) SLC7A11 mRNA levels in tumor tissues with different treatment were detected by qRT-pcr analysis. (e) Representative immunofluorescence staining of total ROS (green) in tumor tissues with different treatment. (f) The morphology of mitochondrial ridges (marked by red arrows) in tumor tissues with different treatment were evaluated by transmission electron microscopy. Each dot represents a mouse. Error bars represent the SEM. ns p >0.05, * p <0.05, ** p <0.01, *** p <0.001.

Article Snippet: After blocking for 15 min with a quickblock buffer (Beyotime, P0235; Shanghai, China), the primary antibody was used upon an overnight incubation at 4°C, including Anti -Interferon γ Rabbit pAb (1:500, Servicebio, GB11107–1), Anti-xCT Rabbit monoclonal antibody (1:1000, Abcam, ab175186), anti-STAT1 Rabbit polyclonal antibody (1:2000, Proteintech 10,144–2-AP), anti-pSTAT1 Rabbit polyclonal antibody (1:1000, Proteintech 28,977–1-AP)and Anti-GAPDH Rabbit monoclonal antibody (1:5000, Abcam, ab181602).

Techniques: Expressing, Western Blot, Binding Assay, Quantitative RT-PCR, Immunofluorescence, Staining, Transmission Assay, Electron Microscopy

Journal: Gut Microbes

Article Title: Synergistic activity of Enterococcus Faeciu m-induced ferroptosis via expansion of IFN-γ + CD8 + T cell population in advanced hepatocellular carcinoma treated with sorafenib

doi: 10.1080/19490976.2024.2410474

Figure Lengend Snippet: Schematic illustration of EPS secretion by efm to induce IFN-γ + CD8 + T cells to promote ferroptosis induced by sorafenib in hepatocellular carcinoma. The EPS secreted by efm in the intestine enters the tumor microenvironment of HCC and prompts IFN-γ + CD8 + T cells to secrete ifn-γ. ifn-γ enters HCC cells to activate the JAK-STAT1 signaling pathway and produce phosphorylated STAT1, which acts as a transcription factor to inhibit the transcription of SLC7A11, thereby increasing intracellular ROS and inducing ferroptosis in HCC cells.

Article Snippet: After blocking for 15 min with a quickblock buffer (Beyotime, P0235; Shanghai, China), the primary antibody was used upon an overnight incubation at 4°C, including Anti -Interferon γ Rabbit pAb (1:500, Servicebio, GB11107–1), Anti-xCT Rabbit monoclonal antibody (1:1000, Abcam, ab175186), anti-STAT1 Rabbit polyclonal antibody (1:2000, Proteintech 10,144–2-AP), anti-pSTAT1 Rabbit polyclonal antibody (1:1000, Proteintech 28,977–1-AP)and Anti-GAPDH Rabbit monoclonal antibody (1:5000, Abcam, ab181602).

Techniques:

Journal: PLoS ONE

Article Title: Interferon-Alpha Triggers B Cell Effector 1 (Be1) Commitment

doi: 10.1371/journal.pone.0019366

Figure Lengend Snippet: In 1A , IFNAR1 and IFNAR2 expression was analyzed in the CD3 − CD19 + CD27 + and CD3 − CD19 + CD27 − lymphocyte gates. A representative staining profile is shown in the left-hand graph. The right-hand graph represents corrected mean fluorescence intensities (MFI) of IFNAR1 and IFNAR2 after substraction of MFI values obtained in isotype control in naive (CD27 − ) and memory (CD27 + ) B cell subsets. The results correspond to the mean ± SEM of the values obtained with cells from 6 healthy donors. 1B shows the purity of B cell preparations (see methods). In 1C , purified B cells were activated with IFN-α for 1 h then lysed. Western blotting was performed on whole-cell lysates by using anti-phospho-STAT2, and the membranes were reprobed with anti-STAT2. The data shown in 1C are representative of 2 independent experiments. 1D, left panel : B cells were activated for 1 hour with IFN-α. Western blotting was performed on whole-cell lysates by using anti-phospho-STAT1 or anti-phospho-STAT4. The membranes were then reprobed with anti-STAT1 or anti-STAT4. The data shown in 1D , left panel are representative of 2 independent experiments. 1D, right panel : B cells were activated with IFN-α for 1 hour or left untreated. They were then fixed, permeabilized, and stained with anti-STAT4 or anti-STAT1 (green) plus propidium iodide (nuclear staining, red). Nuclear translocation was examined by confocal microscopy. Yellow spots indicate nuclear STAT. Similar results were obtained in three other experiments. In 1E , the kinetics of STAT1 and STAT4 phosphorylation was analyzed in B cells by flow cytometry with phospho-STAT-specific antibodies. A representative staining profile is shown in the left-hand graph. The right-hand graph represents corrected mean fluorescence intensities (MFI), after subtraction of MFI values obtained in isotype controls, of phospho-STAT1 and phospho-STAT4 in B cells. The data shown in 1E are the mean ± SEM for cells from 4 healthy donors.

Article Snippet: Proteins were fractionated by 8% sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) in reducing conditions, then transferred to polyvinylidene difluoride filters (PVDF, Boehringer Mannheim) and probed with rabbit polyclonal anti-pSTAT4 IgG (Zymed), rabbit polyclonal anti-pSTAT2 IgG (Upstate Biotechnology) or rabbit polyclonal anti-pSTAT1 IgG (New England Biolabs), followed by the secondary antibody (horseradish peroxidase (HRP)-conjugated goat anti-mouse or anti-rabbit IgG (Jackson Immunoresearch).

Techniques: Expressing, Staining, Fluorescence, Purification, Western Blot, Translocation Assay, Confocal Microscopy, Flow Cytometry