Journal: Cancer biology & therapy

Article Title: Lack of AKT activation in lung cancer cells with EGFR mutation is a novel marker of cetuximab sensitivity.

doi: 10.4161/cbt.19238

Figure Lengend Snippet: Figure 2. Phosphorylation of EGFR, ERK, AKT and STAT3, and expression and mutation status of PTEN in EGFR mutant and wild-type NSCLC cell lines. (A–D) Cells were incubated with/without serum for 24 h, and immunoblotting was done with antibod- ies targeting phosphorylated (p) or total EGFR, ERK and AKT. Densitometric analysis was performed with TotaLab software (Nonlinear Dynamics), and the rela- tive pEGFR/EGFR, pERK/ERK, pAKT/AKT, pSTAT3/STAT3 ratios are indicated as bar graphs under each blot. Experiments were repeated three times and represen- tative results are shown. (E) PTEN protein status in each cell line was determined by immunoblotting in the presence or absence of serum. The β-actin was used as loading control. Experiments were repeated three times and representative results are shown. The muta- tion status of PTEN in each cell line is also indicated. WT, wild type.

Article Snippet: The following primary antibodies were used: pEGFR (Tyr 1068), pAKT (Ser 473), AKT, pSTAT3 (Tyr 705), STAT3, PTEN (Cell Signaling, MA), EGFR (1005) (Santa Cruz), β-ACTIN (Sigma), pERK and ERK (Promega).

Techniques: Phospho-proteomics, Expressing, Mutagenesis, Incubation, Western Blot, Software, Control

Journal: Virology Journal

Article Title: Influenza a virus regulates interferon signaling and its associated genes; MxA and STAT3 by cellular miR-141 to ensure viral replication

doi: 10.1186/s12985-023-02146-4

Figure Lengend Snippet: Oligonucleotides sequences used for quantifying miR-141 and mRNA of indicated genes

Article Snippet: For staining MxA and STAT3, the primary antibodies; rabbit polyclonal anti-MxA (Novus Biologicals, NBP132905) and mouse monoclonal anti-STAT3 (Abcam, ab119352) were used.

Techniques:

Journal: Virology Journal

Article Title: Influenza a virus regulates interferon signaling and its associated genes; MxA and STAT3 by cellular miR-141 to ensure viral replication

doi: 10.1186/s12985-023-02146-4

Figure Lengend Snippet: The correlation between miR-141 level and the expression profile of MxA and STAT3 in infected A549 cells. (A) Quantification of steady-state miR-141 in infected A549 cells with MOI of 0.5 and transfected with either pre-miR-141 or miR-141 inhibitor compared with noninfected cells (control) using qRT-PCR. (B) Relative gene expression of MxA and STAT3 in infected A549 cells transfected with either specific inhibitor against miR-141 or pre- miR-141 compared with control-transfected cells using qRT-PCR. Error bars indicate the STD of three independent experiments. Student two-tailed t -test used for statistical analysis, (*) indicates P-values ≤ 0.05, and (**) indicates P ≤ 0.01. (C) Flow cytometric assay quantifies the kinetic proteins expression profile of MxA (in blue dots) and STAT3 (in red dots) in infected and transfected A549 cells compared with control cells. (D) Western blot analysis reveals the protein expression level of MxA and STAT3 in infected and transfected cells compared to control cells, β-actin expression profile severed as an internal control

Article Snippet: For staining MxA and STAT3, the primary antibodies; rabbit polyclonal anti-MxA (Novus Biologicals, NBP132905) and mouse monoclonal anti-STAT3 (Abcam, ab119352) were used.

Techniques: Expressing, Infection, Transfection, Control, Quantitative RT-PCR, Gene Expression, Two Tailed Test, Flow Cytometry, Western Blot

Journal: Virology Journal

Article Title: Influenza a virus regulates interferon signaling and its associated genes; MxA and STAT3 by cellular miR-141 to ensure viral replication

doi: 10.1186/s12985-023-02146-4

Figure Lengend Snippet: Quantification analysis of miR-141, MxA, and STAT3 in transfected and infected A549 cells

Article Snippet: For staining MxA and STAT3, the primary antibodies; rabbit polyclonal anti-MxA (Novus Biologicals, NBP132905) and mouse monoclonal anti-STAT3 (Abcam, ab119352) were used.

Techniques: Transfection, Infection, Expressing, Control

Journal: Photochemical & photobiological sciences : Official journal of the European Photochemistry Association and the European Society for Photobiology

Article Title: 310 nm UV-LEDs attenuate imiquimod-induced psoriasis-like skin lesions in C57BL/6 mice and inhibit IL-22-induced STAT3 expression in HaCaT cells.

doi: 10.1039/c9pp00444k

Figure Lengend Snippet: Fig. 1 Effects of 310-nm UV-LED on IL-22-induced STAT3 activation in HaCaT cells (a) Emission spectra of 310 nm UV-LED. (b) UV-LEDs cytotoxicity to HaCaT cells was measured by CCK-8 assays. Cells in 96-well microplates (2 × 104 cells/well) were irradiated with 310 nm UV-LED and 311 nm NBUVB (30, 40, 50, and 60 mJ/cm2) for 24 h (black bars). Cells (2 × 104 cells/well) were also pre-treated with IL-22 (100 ng/mL) for 1 h before radiation (grey bars). Unirradiated cells were the control group. Values are mean±SD of three independent experiments. (c) Cell lysates were prepared, and serine 727 phosphorylation (pS-STAT3) was determined in cells stimulated by IL-22 (100 ng/mL) for 0, 5, 10, 30, 60, 120, and 180 minutes. (d) Cells were pre-treated with IL-22 (100 ng/mL for 1 h and irradiated with UV-LEDs at indicated energy levels for 24 h. Cell lysates were prepared and subjected to western blotting for pS-STAT3, p-ERK, and β-actin.

Article Snippet: Sections were stained with monoclonal antibodies against serine 727 (pS-STAT3) (1:100, ab86430, Abcam) and S100A9 (1:100, NB110-89726, Novus Biologicals) as described previously.

Techniques: Activation Assay, CCK-8 Assay, Irradiation, Control, Phospho-proteomics, Western Blot

Journal: Photochemical & photobiological sciences : Official journal of the European Photochemistry Association and the European Society for Photobiology

Article Title: 310 nm UV-LEDs attenuate imiquimod-induced psoriasis-like skin lesions in C57BL/6 mice and inhibit IL-22-induced STAT3 expression in HaCaT cells.

doi: 10.1039/c9pp00444k

Figure Lengend Snippet: Fig. 7 Irradiation of 310 nm UV-LED suppressed skin inflammation via STAT3/S100A9 in imiquimod-induced mice. (a) Tissue extracts from mouse back skins were prepared and blotted to evaluate pS-STAT3, total STAT3, and S100A9 expression. (b) Relative protein ratios were determined as band intensities with ratios of pS-STAT3 and S100A9 in each irradiation group relative to the control. Significant differences from the control group are *P<0.05, ***P<0.001 versus the control group.

Article Snippet: Sections were stained with monoclonal antibodies against serine 727 (pS-STAT3) (1:100, ab86430, Abcam) and S100A9 (1:100, NB110-89726, Novus Biologicals) as described previously.

Techniques: Irradiation, Expressing, Control

Journal: Rheumatology (Sunnyvale, Calif.)

Article Title: Monosodium Urate and Tumor Necrosis Factor-α Increase Apoptosis in Human Chondrocyte Cultures

doi: 10.4172/2161-1149.1000113

Figure Lengend Snippet: Western blotting for p38 kinase, JNK and STAT3. Human OA chondrocytes were grown to hyperdensity as previously described [ , ] and treated with rhTNF-α (10ng/ml) or not (C) for up to 60 min. Cytosolic protein lysate from the 1, 5, 30 and 60 min incubations were electrophoresed as previously described and Western blotting performed . Panels A and B: Western blots were produced as previously described using anti-phospho-p-38 MAPK antibody or an anti-p38 MAPK antibody; ( Panel A ), anti-phospho-SAPK p54 (JNK 2) antibody/anti-phospho-SAPK p46 (JNK 1) antibody or ( Panel B ) anti-SAPK p54 (JNK 2)/anti-SAPK p46 (JNK 1) antibody. Panel C: A Western blot was produced with the specific anti-U-STAT3 antibody or anti-phospho-STAT3 antibody as indicated in Materials and Methods. The bands migrating faster than the phospho-STAT3 standard have retained reactivity with anti-phospho-STAT3, but have not been fully characterized. These bands may represent degradation products of phospho-STAT3. The fastest migrating band reacting with anti-phospho-STAT3 was also present in the control (C) group.

Article Snippet: For immunohistochemical detection of STAT3, an anti-STAT3 monoclonal antibody that detects endogenous human, mouse and rat unphosphorylated STAT3 (U-STAT3) and an affinity-purified rabbit anti-STAT3 antibody that detects STAT3 phosphorylated at Y705 (p-STAT3) were obtained from R and D Systems.

Techniques: Western Blot, Produced, Control

Journal: Rheumatology (Sunnyvale, Calif.)

Article Title: Monosodium Urate and Tumor Necrosis Factor-α Increase Apoptosis in Human Chondrocyte Cultures

doi: 10.4172/2161-1149.1000113

Figure Lengend Snippet: U-STAT3 and p-STAT3 in normal juvenile chondrocyte pellet cultures (N-JHu-C) and pellet cultures initiated from MSCs (BMD-MSC-C).

Article Snippet: For immunohistochemical detection of STAT3, an anti-STAT3 monoclonal antibody that detects endogenous human, mouse and rat unphosphorylated STAT3 (U-STAT3) and an affinity-purified rabbit anti-STAT3 antibody that detects STAT3 phosphorylated at Y705 (p-STAT3) were obtained from R and D Systems.

Techniques:

Journal: BMC Medicine

Article Title: ISG15-driven immune modulation and tumor progression in breast cancer metastasis: insights from single-cell and spatial transcriptomics

doi: 10.1186/s12916-025-04614-w

Figure Lengend Snippet: Transcriptomic analysis of in situ tumor tissues in mouse mammary gland cancer. Note: A The volcano plot of differentially expressed genes in in situ tumor tissues of mouse mammary gland cancer after ISG15 silencing, with red dots representing upregulated genes and blue dots representing downregulated genes; B heatmap depicting the expression levels of differentially expressed genes in the sh-NC group (6 cases) and sh-ISG15 group (6 cases), with the left dendrogram clustering genes based on expression levels, the color bar on the right representing expression levels (red indicating upregulated genes and blue indicating downregulated genes), and the histogram above indicating blue for non-metastatic group and red for metastatic group; C GO analysis of differentially expressed genes, with the color bar on the right representing enrichment scores for different functional categories; D KEGG analysis of differentially expressed genes, with the color bar on the right representing enrichment scores for different pathways; E expression levels of PD-L1 in in situ tumor tissues of mouse mammary gland cancer after ISG15 silencing, with 6 mice in each group; F binding sites of transcriptional activation factor STAT3

Article Snippet: The membrane was blocked with 5% skim milk at room temperature (RT) for 1 h, then incubated overnight at 4 °C with primary antibodies against ISG15 (1:1000, ab308219), NESTIN (1:1000, ab221660), OCT4 (1:10,000, ab200834), SOX2 (1:1500, ab92494), STAT3 (1:1500, ab68153), p-JAK2 (1:1500, ab32101), JAK2 (1:5000, ab108596), PD-L1 (1:1000, ab213480), Arg (1:1000, ab203490), CD206 (1:1200, ab64693), CD163 (1:1000, ab182422), CD86 (1:1200, ab220188), and GAPDH (1:2500, ab9485) as the internal reference (all from Abcam, Cambridge, UK), as well as p-STAT3 (1:1000, NB100-82213, Novus Biologicals).

Techniques: In Situ, Expressing, Functional Assay, Binding Assay, Activation Assay

Journal: BMC Medicine

Article Title: ISG15-driven immune modulation and tumor progression in breast cancer metastasis: insights from single-cell and spatial transcriptomics

doi: 10.1186/s12916-025-04614-w

Figure Lengend Snippet: Influence of ISG15 in CSCs on T cell activation and its potential molecular mechanisms. Note: A Flow cytometry analysis of Ki-67 to determine the proportion of proliferating CD8 + T cells; B flow cytometry analysis of IFN-γ to determine the proportion of activated CD8 + T cells; C western blot analysis of JAK-STAT signaling pathway-related proteins and PD-L1 expression in co-culture systems of different groups; D dual-luciferase assay to examine the impact of ISG15 on PD-L1 promoter activity; E the performance of ChIP assay to examine the enrichment of p-STAT3 on the PD-L1 promoter in 4T1-S cells (left panel) and CSCs isolated from mouse tumor models (right panel); F the utilization of flow cytometry for the detection of Ki-67 expression in T cells; G The application of flow cytometry for the detection of IFN-γ expression in T cells; H the use of flow cytometry for the detection of Ki-67 expression in T cells; I the use of flow cytometry for the detection of IFN-γ expression in T cells; J the performance of western blot for the analysis of PD-L1 expression in each group co-culture system; K schematic representation of the molecular mechanism of ISG15 in T cell activation in CSCs. * indicates statistical significance ( P < 0.05) between the two groups. All cellular experiments were performed in triplicate

Article Snippet: The membrane was blocked with 5% skim milk at room temperature (RT) for 1 h, then incubated overnight at 4 °C with primary antibodies against ISG15 (1:1000, ab308219), NESTIN (1:1000, ab221660), OCT4 (1:10,000, ab200834), SOX2 (1:1500, ab92494), STAT3 (1:1500, ab68153), p-JAK2 (1:1500, ab32101), JAK2 (1:5000, ab108596), PD-L1 (1:1000, ab213480), Arg (1:1000, ab203490), CD206 (1:1200, ab64693), CD163 (1:1000, ab182422), CD86 (1:1200, ab220188), and GAPDH (1:2500, ab9485) as the internal reference (all from Abcam, Cambridge, UK), as well as p-STAT3 (1:1000, NB100-82213, Novus Biologicals).

Techniques: Activation Assay, Flow Cytometry, Western Blot, Expressing, Co-Culture Assay, Luciferase, Activity Assay, Isolation

Journal: Journal of Pharmaceutical Analysis

Article Title: CXCL8/SDC1 axis mediates tumor stem cell interactions to drive remote transfer in thyroid cancer

doi: 10.1016/j.jpha.2025.101354

Figure Lengend Snippet: Impact of the Janus kinase-signal transducer and activator of transcription (JAK-STAT) signaling pathway on tumor stem cell self-renewal and migration invasion. (A, B) Western blot analysis of JAK-STAT and nuclear factor kappa B (NF-κB) signaling pathway-related proteins: protein expression levels of phosphorylated-Janus kinase 2 (p-JAK2), JAK2, phosphorylated signal transducer and activator of transcription (p-STAT3), STAT3, NF-κB, and phosphorylated NF-κB (p-NF-κB) in FTC238-S cells co-cultured with different modified myeloid‑derived suppressor cells (MDSCs) (A), and protein levels of p-JAK2, JAK2, p-STAT3, and STAT3 in FTC238-S cells treated with anti-immunoglobulin G (anti-IgG) or anti-C-X-C motif chemokine ligand 8 (anti-CXCL8) antibodies (B). (C) Schematic diagram of monocyte treatment and co-culturing with FTC238-S cells. (D) Western blot analysis of JAK-STAT signaling pathway-related proteins in FTC238-S cells from each group. (E) Protein expression levels of stemness markers neuroepithelial stem cell protein (NESTIN), octamer-binding transcription factor 4 (OCT4), and SRY-box transcription factor 2 (SOX2) in FTC238-S cells from each group examined by Western blot. (F) Cell sphere formation assay assessing the sphere formation capability of FTC238-S cells in co-culture systems of each group. (G) Clonogenic assay evaluating the clonogenic capacity of FTC238-S cells in co-culture systems of each group. (H) Cell Counting Kit-8 (CCK-8) assay measuring the proliferation ability of FTC238-S cells in co-culture systems of each group. (I) Transwell assay determining the migration and invasion capability of FTC238-S cells in co-culture systems of each group, ∗ P < 0.05 compared between the two groups, and all cell experiments were repeated three times. M_oe-NC + S_sh-NC: FTC238-S cells transfected with short hairpin RNA-negative control (sh-NC) and treated with conditioned medium from monocytes transfected with oe-NC; M_oe-CXCL8 + S_sh-NC: FTC238-S cells transfected with sh-NC and treated with conditioned medium from monocytes transfected with oe-CXCL8; M_oe-CXCL8 + S_sh-SDC1: FTC238-S cells transfected with short hairpin RNA-SDC1 (sh-SDC1) and treated with conditioned medium from monocytes transfected with oe-CXCL8; M_oe-CXCL8 + S_DMSO: FTC238-S cells treated with conditioned medium from monocytes transfected with oe-CXCL8 and supplemented with an equal amount of dimethyl sulfoxide (DMSO); M_oe-CXCL8 + S_SD_1008: FTC238-S cells treated with conditioned medium from monocytes transfected with oe-CXCL8 and supplemented with SD-1008; THP-1 cells: Tohoku Hospital Pediatrics-1 cells; OD: optical density.

Article Snippet: Proteins were transferred to a polyvinylidene difluoride (PVDF) membrane and blocked with 5% skim milk at room temperature for 1 h. The PVDF membrane was incubated overnight at 4 °C with primary antibodies against CXCL8 (1:1000, ab235584, Abcam, Cambridge, UK), SDC1 (1:2000, ab128936, Abcam, Cambridge, UK), NESTIN (1:100, ab105389, Abcam, Cambridge, UK), OCT4 (1:10000, ab200834, Abcam, Cambridge, UK), SRY-Box transcription factor 2 (SOX2, 1:1500, ab92494, Abcam, Cambridge, UK), phosphorylated STAT3 (p-STAT3) (1:1000, NB100-82213, Novus Biologicals, Centennial, CO, USA), STAT3 (1:1000, MAB1799, Novus Biologicals, Centennial, CO, USA), phosphorylated-JAK2 (p-JAK2) (1:1500, ab32101, Abcam, Cambridge, UK), JAK2 (1:1000, ab108596, Abcam, Cambridge, UK), nuclear factor kappa B (NF-κB, 1:1000, 8242T, CST, Boston, MA, USA), phosphorylated NF-κB (p-NF-κB) (1:1000, 3033T, CST, Boston, MA, USA), glyceraldehyde 3-phosphate dehydrogenase (GAPDH) (1:2500, ab9485, Abcam, Cambridge, UK), and Tubulin (1:5000, ab7291, Abcam, Cambridge, UK).

Techniques: Migration, Western Blot, Expressing, Cell Culture, Modification, Binding Assay, Tube Formation Assay, Co-Culture Assay, Clonogenic Assay, Cell Counting, CCK-8 Assay, Transwell Assay, Transfection, shRNA, Negative Control

Journal: Journal of Pharmaceutical Analysis

Article Title: CXCL8/SDC1 axis mediates tumor stem cell interactions to drive remote transfer in thyroid cancer

doi: 10.1016/j.jpha.2025.101354

Figure Lengend Snippet: Impact of the C-X-C motif chemokine ligand 8/syndecan-1 (CXCL8/SDC1) axis on tumor initiation, growth, and metastasis of thyroid cancer (THCA) stem cells i n Vivo . (A) Diagram of the in vivo animal experiment protocol, with the green syringe representing anti-immunoglobulin G (anti-IgG) and anti-CXCL8 treatments. (B) Western blot analysis of Janus kinase-signal transducer and activator of transcription (JAK-STAT) signaling pathway-related proteins in subcutaneous tumor tissues of nude mice from each group. (C) Gross anatomy of subcutaneous transplant tumors in nude mice (left) and corresponding weight statistics (right). (D) Images of primary tumors in nude mice from each group (left) and diameter analysis (right). (E) Statistical analysis of axillary lymph node (LN) metastasis in subcutaneous transplant tumors of nude mice from each group. (F) Hematoxylin and eosin (H&E) staining to assess bone metastasis of primary tumors in nude mice from each group. (G) Images of lung metastasis in nude mice from each group (left) and diameter analysis (right). (H) H&E staining to evaluate lung metastasis in nude mice from each group. ∗ P < 0.05 compared between two groups, ∗∗ P < 0.01 with 6 nude mice in each group. p-JAK2: phosphorylated-JAK2; p-STAT3: phosphorylated STAT3; GAPDH: glyceraldehyde 3-phosphate dehydrogenase; S_sh-NC: FTC238-S cells transfected with short hairpin RNA-negative control (sh-NC); S_sh-SDC1: FTC238-S cells transfected with short hairpin RNA-SDC1 (sh-SDC1).

Article Snippet: Proteins were transferred to a polyvinylidene difluoride (PVDF) membrane and blocked with 5% skim milk at room temperature for 1 h. The PVDF membrane was incubated overnight at 4 °C with primary antibodies against CXCL8 (1:1000, ab235584, Abcam, Cambridge, UK), SDC1 (1:2000, ab128936, Abcam, Cambridge, UK), NESTIN (1:100, ab105389, Abcam, Cambridge, UK), OCT4 (1:10000, ab200834, Abcam, Cambridge, UK), SRY-Box transcription factor 2 (SOX2, 1:1500, ab92494, Abcam, Cambridge, UK), phosphorylated STAT3 (p-STAT3) (1:1000, NB100-82213, Novus Biologicals, Centennial, CO, USA), STAT3 (1:1000, MAB1799, Novus Biologicals, Centennial, CO, USA), phosphorylated-JAK2 (p-JAK2) (1:1500, ab32101, Abcam, Cambridge, UK), JAK2 (1:1000, ab108596, Abcam, Cambridge, UK), nuclear factor kappa B (NF-κB, 1:1000, 8242T, CST, Boston, MA, USA), phosphorylated NF-κB (p-NF-κB) (1:1000, 3033T, CST, Boston, MA, USA), glyceraldehyde 3-phosphate dehydrogenase (GAPDH) (1:2500, ab9485, Abcam, Cambridge, UK), and Tubulin (1:5000, ab7291, Abcam, Cambridge, UK).

Techniques: In Vivo, Western Blot, Staining, Transfection, shRNA, Negative Control

Journal: Journal of Pharmaceutical Analysis

Article Title: CXCL8/SDC1 axis mediates tumor stem cell interactions to drive remote transfer in thyroid cancer

doi: 10.1016/j.jpha.2025.101354

Figure Lengend Snippet: Schematic diagram of the molecular mechanism by which monocytes promote self-renewal, migration, and invasion of thyroid cancer (THCA) stem cells through the C-X-C motif chemokine ligand 8/syndecan-1 (CXCL8/SDC1) axis. JAK2: Janus kinase 2; STAT3: signal transducer and activator of transcription 3.

Article Snippet: Proteins were transferred to a polyvinylidene difluoride (PVDF) membrane and blocked with 5% skim milk at room temperature for 1 h. The PVDF membrane was incubated overnight at 4 °C with primary antibodies against CXCL8 (1:1000, ab235584, Abcam, Cambridge, UK), SDC1 (1:2000, ab128936, Abcam, Cambridge, UK), NESTIN (1:100, ab105389, Abcam, Cambridge, UK), OCT4 (1:10000, ab200834, Abcam, Cambridge, UK), SRY-Box transcription factor 2 (SOX2, 1:1500, ab92494, Abcam, Cambridge, UK), phosphorylated STAT3 (p-STAT3) (1:1000, NB100-82213, Novus Biologicals, Centennial, CO, USA), STAT3 (1:1000, MAB1799, Novus Biologicals, Centennial, CO, USA), phosphorylated-JAK2 (p-JAK2) (1:1500, ab32101, Abcam, Cambridge, UK), JAK2 (1:1000, ab108596, Abcam, Cambridge, UK), nuclear factor kappa B (NF-κB, 1:1000, 8242T, CST, Boston, MA, USA), phosphorylated NF-κB (p-NF-κB) (1:1000, 3033T, CST, Boston, MA, USA), glyceraldehyde 3-phosphate dehydrogenase (GAPDH) (1:2500, ab9485, Abcam, Cambridge, UK), and Tubulin (1:5000, ab7291, Abcam, Cambridge, UK).

Techniques: Migration

Journal: Signal Transduction and Targeted Therapy

Article Title: Bioengineered iPSC-derived human macrophages with increased angiotensin-converting enzyme (ACE) expression suppress solid tumor growth

doi: 10.1038/s41392-026-02650-3

Figure Lengend Snippet: Determination of M1 vs. M2 polarization in iMac. a Pro-inflammatory M1 cytokines IL-1β, TNFα, IL-6, and IL-8 measured by ELISA. b Anti-inflammatory M2 cytokines IL-4, IL-10, IL-13, CCL5, CCL17, and CCL20 measured by ELISA. For a , b iMac were cultured in 6-well plates (3 × 10 6 cells/well). To challenge with tumor factors, iMac were co-cultured with 50% fresh supernatant from SK-MEL-28 cells every 24 h for 3 days. Supernatants were collected during the final 24 h for cytokine measurement. c Activation (phosphorylation) of NF-kB p65, STAT1, STAT3, and STAT6 in iMac was determined using Western blotting after conditioning with melanoma (SK-MEL-28) supernatant for 72 h. Protein band intensity was quantified using Odyssey version 3.0 software. After adjusting for loading using the β-actin band, data are presented as fold change to the iMac (Dox-) group, which is set as 1. For a – c , one-way ANOVA with Bonferroni’s correction for multiple comparisons was used to analyze group comparisons. Data are presented as means ± SD. Each experiment was performed three times, with triplicate samples each time. * p < 0.05, *** p < 0.001, and **** p < 0.0001

Article Snippet: The polyvinylidene difluoride (PVDF) membranes were incubated with specific primary antibodies against ACE (R&D Systems, MAB9291, 1:1,000), GAPDH (Sigma-Aldrich, SAB5600208, 1:2,000), β-actin (Sigma-Aldrich, A3854; 1:1000), phosphorylated NF-kB p65 (Novus, NB100-82086, 1:500), phosphorylated STAT1 (R&D Systems, AF2894, 1 μg/ml), phosphorylated STAT3 (Novus, NBP2-24463, 0.5 μg/ml), or phosphorylated STAT6 (Millipore, 06-937, 1:1000).

Techniques: Enzyme-linked Immunosorbent Assay, Cell Culture, Activation Assay, Phospho-proteomics, Western Blot, Software

Journal: eLife

Article Title: Unsupervised machine learning reveals risk stratifying glioblastoma tumor cells

doi: 10.7554/eLife.56879

Figure Lengend Snippet:

Article Snippet: Antibody , Anti-p-STAT3 (Y705) (mouse-monoclonal) , Fluidigm , RRID: AB_2811100 Cat# 3158005A Clone: 4/P-STAT3 , MC (1:100).

Techniques: Isolation, Software