phosphor stat3 rabbit mab  (Cell Signaling Technology Inc)

Journal: NPJ Precision Oncology

Article Title: RICH1 enhances pro-inflammatory TAM infiltration in breast cancer via promoting TRIM21-mediated ubiquitination of RhoA and inhibiting STAT3 phosphorylation

doi: 10.1038/s41698-025-01252-6

Figure Lengend Snippet: A The top lists RICH1 binding proteins in MASS analysis (IP RICH1 in RICH1 overexpressed 4T1 cells), including RICH1 and its functionally related proteins, and innate immune-related TRIM21. B The protein-protein interaction network between RICH1, TRIM21, and RhoA-STAT3 identified by STRING database. RICH1, as a Rho GTPase activating protein, interacts with RhoA, and STAT3 was reported interacting with RhoA, TRIM21, and IFNG in previous studies. C Western blotting after Co-IP analysis for confirmation of interaction between RICH1-TRIM21 in 4T1 cells. D Detection of main above molecules after RICH1 overexpression and knockout by Western blotting. E IHC staining and score for RICH1, RhoA-STAT3, and IFN-γ of same fields on serial paraffin sections of 4T1 transplanted tumors. All the experiments were repeated three times independently with similar results. Abbreviation: Co-IP co-immunoprecipitation, MASS mass spectrometry, ARHGAP17 gene symbol for RICH1, HC heavy chain, IHC immunohistochemistry.

Article Snippet: Antibodies against STAT3 (#9139, dilution 1:500), phosphor-STAT3 Tyr705 (#9145, dilution 1:1000), Cleaved Caspase‐8 (#8592, dilution 1:500), and Pan-Cytokeratin (#4545, dilution 1:200) were purchased from Cell Signaling Technology.

Techniques: Binding Assay, Western Blot, Co-Immunoprecipitation Assay, Over Expression, Knock-Out, Immunohistochemistry, Immunoprecipitation, Mass Spectrometry

Journal: NPJ Precision Oncology

Article Title: RICH1 enhances pro-inflammatory TAM infiltration in breast cancer via promoting TRIM21-mediated ubiquitination of RhoA and inhibiting STAT3 phosphorylation

doi: 10.1038/s41698-025-01252-6

Figure Lengend Snippet: A Detection of RhoA expression after CHX (100 μg/ml) contacting HC11 cells with gradient time by Western blotting and quantification of RhoA protein levels determined using Image J, normalized to GAPDH. B Detection of RhoA expression after MG132 (5 μM) contacting 4T1 and HC11 cells with 8 h by Western blotting. C The ubiquitination of RhoA was examined by IP of RhoA and Western blotting with anti-ubiquitin antibody after ubiquitin-HA was transfected into RICH1 overexpression and knockout cells. D Western blotting after Co-IP analysis for confirmation of interaction between RhoA-TRIM21 in 4T1 cells with RICH1 overexpression. E Detection of the ubiquitination of RhoA and protein levels of RICH1, TRIM21, and RhoA-STAT3 by Western blotting after inhibiting the expression of TRIM21 in RICH1 overexpressed 4T1 cells. F , G Detection of the active form of RhoA (RhoA-GTP) in above cells by the pull-down assay. All the experiments were repeated three times independently with similar results. H , I The visualized image of molecular docking between RICH1 (yellow), TRIM21 (blue), and RhoA (green). J The volume and weight of the tumors compared in the mice injected with 4T1-RICH1 OE shTRIM21/4T1-RICH1 OE. K The proportions of CD86 + subpopulations in CD45 + CD11b + F4/80 + cells of TME in the above tissues by flow cytometry analysis. Abbreviation: CHX cycloheximide, Co-IP co-immunoprecipitation, HC heavy chain. P values in ( J and K ) were calculated using unpaired two-tailed Student’s t tests. * P < 0.05; *** P < 0.001.

Article Snippet: Antibodies against STAT3 (#9139, dilution 1:500), phosphor-STAT3 Tyr705 (#9145, dilution 1:1000), Cleaved Caspase‐8 (#8592, dilution 1:500), and Pan-Cytokeratin (#4545, dilution 1:200) were purchased from Cell Signaling Technology.

Techniques: Expressing, Western Blot, Ubiquitin Proteomics, Transfection, Over Expression, Knock-Out, Co-Immunoprecipitation Assay, Pull Down Assay, Injection, Flow Cytometry, Immunoprecipitation, Two Tailed Test

Journal: Discover Oncology

Article Title: The involvement of exosomes in the tumorigenicity of breast cancer cell lines through the crosstalk between STAT3, Notch, and Wnt signaling pathways

doi: 10.1007/s12672-025-03334-0

Figure Lengend Snippet: Graphical Abstract. A 1. Exosomes isolation from serum, 2. Purification with ultracentrifuge. B Exosomes characterization, 3. Scanning electron microscope and 4. Western blotting then 5. Labeling with PKH47 dye followed by assessment of 6. Uptake efficiency by three cell lines C 7. STAT3 pathway inhibition using AG490. D 8. Crosstalk assessment between STAT3, WNT and NOTCH signalling

Article Snippet: Using the western blotting technique, the pan-STAT3 (STAT3 mAb, Wuhan Fine Biotech Co., Ltd., China), phosphor-STAT3 (Tyr705-STAT3 mAb, Cell Signaling Technology, USA) proteins, and GAPDH protein (Novus Biologicals, LLC, USA) as internal control, were determined to assess the activity of STAT3 signaling.

Techniques: Isolation, Purification, Microscopy, Western Blot, Labeling, Inhibition

Journal: Discover Oncology

Article Title: The involvement of exosomes in the tumorigenicity of breast cancer cell lines through the crosstalk between STAT3, Notch, and Wnt signaling pathways

doi: 10.1007/s12672-025-03334-0

Figure Lengend Snippet: A Representative cropped images of western blots before (-) and after (+) STAT3 pathway inhibition with AG490 inhibitor were analyzed using a Gel Dox XR + Gel Documentation System. System (Bio-Rad). Original blot was presented in supplementary Fig. 2. B Plots showing the inhibitory effect (%) after 48 h of incubation. C Plots of the Lane Normalization Factor (LNF) for each cell line

Article Snippet: Using the western blotting technique, the pan-STAT3 (STAT3 mAb, Wuhan Fine Biotech Co., Ltd., China), phosphor-STAT3 (Tyr705-STAT3 mAb, Cell Signaling Technology, USA) proteins, and GAPDH protein (Novus Biologicals, LLC, USA) as internal control, were determined to assess the activity of STAT3 signaling.

Techniques: Western Blot, Inhibition, Incubation

Journal: Discover Oncology

Article Title: The involvement of exosomes in the tumorigenicity of breast cancer cell lines through the crosstalk between STAT3, Notch, and Wnt signaling pathways

doi: 10.1007/s12672-025-03334-0

Figure Lengend Snippet: Morphological alterations in A TNBC cell lines, MDA-MB-231, HCC1806, and B NTNBC cell line, MCF7, after EXO and/or DOXO treatment for 48 h before and after STAT3 pathway inhibition

Article Snippet: Using the western blotting technique, the pan-STAT3 (STAT3 mAb, Wuhan Fine Biotech Co., Ltd., China), phosphor-STAT3 (Tyr705-STAT3 mAb, Cell Signaling Technology, USA) proteins, and GAPDH protein (Novus Biologicals, LLC, USA) as internal control, were determined to assess the activity of STAT3 signaling.

Techniques: Inhibition

Journal: Discover Oncology

Article Title: The involvement of exosomes in the tumorigenicity of breast cancer cell lines through the crosstalk between STAT3, Notch, and Wnt signaling pathways

doi: 10.1007/s12672-025-03334-0

Figure Lengend Snippet: Microscopy images of the migration assay under an inverted microscope after EXO and/or DOXO treatment for 48 h before and after STAT3 pathway inhibition in A TNBC cell lines, MDA-MB-231, HCC1806, and B TNBC cell line, MCF7. Three replicas were used for each experiment and analysis was done using ImagJ software

Article Snippet: Using the western blotting technique, the pan-STAT3 (STAT3 mAb, Wuhan Fine Biotech Co., Ltd., China), phosphor-STAT3 (Tyr705-STAT3 mAb, Cell Signaling Technology, USA) proteins, and GAPDH protein (Novus Biologicals, LLC, USA) as internal control, were determined to assess the activity of STAT3 signaling.

Techniques: Microscopy, Migration, Inverted Microscopy, Inhibition, Software

Journal: Discover Oncology

Article Title: The involvement of exosomes in the tumorigenicity of breast cancer cell lines through the crosstalk between STAT3, Notch, and Wnt signaling pathways

doi: 10.1007/s12672-025-03334-0

Figure Lengend Snippet: Colony formation assay in ( A ) TNBC cell lines, MDA-MB-231, HCC1806, and ( B ) NTNBC cell line, MCF7, after EXO and/or DOXO treatment for 48 h before and after STAT3 pathway inhibition. Three replicas were used for each experiment and analysis was done using ImagJ software

Article Snippet: Using the western blotting technique, the pan-STAT3 (STAT3 mAb, Wuhan Fine Biotech Co., Ltd., China), phosphor-STAT3 (Tyr705-STAT3 mAb, Cell Signaling Technology, USA) proteins, and GAPDH protein (Novus Biologicals, LLC, USA) as internal control, were determined to assess the activity of STAT3 signaling.

Techniques: Colony Assay, Inhibition, Software

Journal: Discover Oncology

Article Title: The involvement of exosomes in the tumorigenicity of breast cancer cell lines through the crosstalk between STAT3, Notch, and Wnt signaling pathways

doi: 10.1007/s12672-025-03334-0

Figure Lengend Snippet: Expression analysis of targeted genes in the Wnt signaling pathways in MDA-MB-231, HCC1806, and MCF7 cells after 48 h of EXO and/or DOXO treatment, both before and after STAT3 pathway inhibition. Six replicas were used for each experiment and analyzed using the SPSS program

Article Snippet: Using the western blotting technique, the pan-STAT3 (STAT3 mAb, Wuhan Fine Biotech Co., Ltd., China), phosphor-STAT3 (Tyr705-STAT3 mAb, Cell Signaling Technology, USA) proteins, and GAPDH protein (Novus Biologicals, LLC, USA) as internal control, were determined to assess the activity of STAT3 signaling.

Techniques: Expressing, Protein-Protein interactions, Inhibition

Journal: Discover Oncology

Article Title: The involvement of exosomes in the tumorigenicity of breast cancer cell lines through the crosstalk between STAT3, Notch, and Wnt signaling pathways

doi: 10.1007/s12672-025-03334-0

Figure Lengend Snippet: Expression analysis of targeted genes in the Notch signaling pathways; A )Notch1, Notch2, and JAG1 B HES1 and DLL4, in MDA-MB-231, HCC1806, and MCF7 cells after 48 h of EXO and/or DOXO treatment, both before and after STAT3 pathway inhibition. Six replicas were used for each experiment and analyzed using the SPSS program

Article Snippet: Using the western blotting technique, the pan-STAT3 (STAT3 mAb, Wuhan Fine Biotech Co., Ltd., China), phosphor-STAT3 (Tyr705-STAT3 mAb, Cell Signaling Technology, USA) proteins, and GAPDH protein (Novus Biologicals, LLC, USA) as internal control, were determined to assess the activity of STAT3 signaling.

Techniques: Expressing, Protein-Protein interactions, Inhibition