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Ptyr705 stat3

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PTyr705-STAT3 is an antibody that detects phosphorylation of STAT3 at tyrosine 705. This post-translational modification is crucial for STAT3 activation and plays a key role in signal transduction pathways.

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Lab products found in correlation

Stat3, by Cell Signaling Technology (3 mentions) P jak2 tyr1007 1008, by Cell Signaling Technology (3 mentions) P erk, by Cell Signaling Technology (3 mentions) Total stat3, by Cell Signaling Technology (3 mentions) Pser727 stat3, by Cell Signaling Technology (3 mentions) Cyclin d1, by Cell Signaling Technology (2 mentions) P jak1 tyr1022 1023, by Cell Signaling Technology (2 mentions) P tyk2 tyr1054 1055, by Cell Signaling Technology (2 mentions) Pakt1 ser 473, by Cell Signaling Technology (2 mentions) Pakt1 thr308, by Cell Signaling Technology (2 mentions) β actin, by Merck Group (2 mentions) Junb and cjun, by Santa Cruz Biotechnology (2 mentions) Pgsk3α β, by Cell Signaling Technology (2 mentions) Socs3, by Cell Signaling Technology (2 mentions) Bcl 2, by Santa Cruz Biotechnology (2 mentions) Bcl xl, by Santa Cruz Biotechnology (2 mentions) Adam17, by Merck Group (1 mentions) α smooth muscle actin, by Merck Group (1 mentions) P p65 ser536, by Cell Signaling Technology (1 mentions) Apoptag peroxidase in situ apoptosis detection kit, by Merck Group (1 mentions)

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STAT3 (1075 citations)

16 protocols using ptyr705 stat3

1

Immunohistochemical Analysis of Pancreatic Tissue

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FFPE human pancreatic sections were stained with antibodies against ADAM17 (Sigma) and pTyr705-STAT3 (Cell Signaling Technology). Mouse pancreatic FFPE sections were subjected to immunohistochemistry (IHC) with the following antibodies: ADAM17, α-Smooth muscle actin (αSMA; Sigma), CD45, B220 (BD Biosciences), CD11c, cleaved caspase-3, pThr180/pTyr182-p38 MAPK, pThr202/pTyr204-ERK1/2 MAPK, pTyr705-STAT3, pSer536 p65 nuclear factor-κB (NF-κB), pSer473 AKT (Cell Signaling Technology), CD3, Ly6G, and F4/80 (Abcam). Cellular apoptosis was determined by the terminal deoxynucleotidyl transferase (tdT)-mediated dUDP nick-end labeling (TUNEL) technique using the ApopTag Peroxidase In Situ Apoptosis Detection kit (Millipore). Masson’s trichome staining was performed using standard protocols at Monash Histology Platform. To quantify cellular staining, digital images of photomicrographs (60× high-power fields) were viewed using Image J software. Positive-stained cells were counted manually or staining intensity was quantified using Image J software (n = 20 fields).
Saad M.I., Weng T., Lundy J., Gearing L.J., West A.C., Harpur C.M., Alanazi M., Hodges C., Croagh D., Kumar B., Sagi I., Rose-John S, & Jenkins B.J. (2022). Blockade of the protease ADAM17 ameliorates experimental pancreatitis. Proceedings of the National Academy of Sciences of the United States of America, 119(42), e2213744119.
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2

Niclosamide and TRA-8 mAb Combination

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Niclosamide was purchased from Sigma (St. Louis, MO). Niclosamide for in vitro use was dissolved in DMSO at a 4.8 mM concentration and stored at 4°C until further use. For animal studies, Niclosamide was dissolved in DMSO until a homogeneous suspension was observed at which time Cremophor was added to make a final solution of 25% DMSO and 75% Cremophor. The liquid was slowly inverted to obtain a clear orange solution, which was stored at 4°C. Purified TRA-8 (IgG1) mAb was prepared at the University of Alabama at Birmingham, as described previously, and was provided by Dr. Tong Zhou (32 (link)). IgG1 and isotype-specific IgG1 control antibody were obtained from Southern Biotechnology Associates (Birmingham, AL). ALDEFLUOR kit including DEAB was obtained from StemCell Technologies (Durham, NC). Monoclonal anti- phosphorylated-LRP6, Axin2, cyclin D1, p(Tyr705)-STAT3 and STAT3 were purchased from Cell Signaling Technology, Inc. (Danvers, MA). Monoclonal anti-β-catenin was purchased from BD Biosciences (San Jose, CA). Survivin antibody for Western blots was purchased from Santa Cruz (Santa Cruz, CA).
Londoño-Joshi A.I., Arend R.C., Aristizabal L., Lu W., Samant R.S., Metge B.J., Hidalgo B., Grizzle W.E., Conner M., Forero-Torres A., LoBuglio A.F., Li Y, & Buchsbaum D.J. (2014). Effect of niclosamide on basal-like breast cancers. Molecular cancer therapeutics, 13(4), 800-811.
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3

Therapeutic Antibodies and Mouse Model

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Antibodies against p-Tyr701-STAT1 (product #9167), p-Tyr690-STAT2 (product #88410), p-Tyr705-STAT3 (product #9145), p-Tyr1022/1023-JAK1 (product #3331), p-Tyr1007/1008-JAK2 (product #3776), and p-Tyr1054/1055-Tyk2 (product #9321) were purchased from Cell Signaling Technology, USA. Recombinant human STAT3 was obtained from Abcam, UK (product #ab268982). Protease inhibitors and phosphatase inhibitors were obtained from Merck Life Science, USA. Marine natural products library was built in house in the Laboratory for Marine Resources Biology at Ocean University of China.
Six-week old female NRMI congenitally athymic nude (nu/nu) mice (SPF degree, 17–20 g weight) were purchased from Beijing Vital River Laboratory Animal Technology Co., Ltd.
Li R., Zhou Y., Zhang X., Yang L., Liu J., Wightman S.M., Lv L., Liu Z., Wang C.Y, & Zhao C. (2023). Identification of marine natural product Pretrichodermamide B as a STAT3 inhibitor for efficient anticancer therapy. Marine Life Science & Technology, 5(1), 94-101.
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4

Western Blot Analysis of Signaling Proteins

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Western blot analysis was performed using methods described previously30 (link). Antibodies against the following proteins were used: AKT1, pAKT1 (Thr308), pAKT1 (Ser 473), STAT3, p-Tyr705 STAT3, ERK, pERK, pALK and pGSK3α/β (Cell Signaling Technology, Beverly, MA, USA), JUNB and CJUN (Santa Cruz Biotechnology, Santa Cruz, CA, USA), AKT2 and AKT3 (Millipore, Billerica, Massachusetts, MA, USA). β-Actin (Sigma, St Louis, MO) served as a control for protein load and integrity in all immunoblots.
Atsaves V., Zhang R., Ruder D., Pan Y., Leventaki V., Rassidakis G.Z, & Claret F.X. (2015). Constitutive control of AKT-1 gene expression by JUNB / CJUN in ALK+ anaplastic large cell lymphoma: a novel crosstalk mechanism. Leukemia, 29(11), 2162-2172.
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5

Histone Modification Analysis in Lymphoma

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PBS Lysis Buffer (1% Triton X-100, 1mM DTT, in PBS) followed by 0.2 N HCl solution was used to prepare lysates for histone fraction of lymphoma/B220+ cells. RIPPA buffer was used to prepare whole cell lysates of OCI-LY7 cells. Immunoblot analyses were performed according to standard procedures. Membranes were probed with the indicated primary antibodies: H3K4me1 (Abcam, #ab8895), H3K4me2 (Millipore #07-030), H3K4me3 (Millipore #07-473), total H3 (abcam #ab1791), P-Tyr705-STAT3 (Cell Signaling #9145), total-STAT3 (Cell Signaling #12640), and SOCS3 (Cell Signaling #2932). Enhanced chemiluminescence was used for detection (ECL, Amersham).
Ortega-Molina A., Boss I.W., Canela A., Pan H., Jiang Y., Zhao C., Jiang M., Hu D., Agirre X., Niesvizky I., Lee J.E., Chen H.T., Ennishi D., Scott D.W., Mottok A., Hother C., Liu S., Cao X.J., Tam W., Shaknovich R., Garcia B.A., Gascoyne R.D., Ge K., Shilatifard A., Elemento O., Nussenzweig A., Melnick A.M, & Wendel H.G. (2015). The histone lysine methyltransferase KMT2D sustains a gene expression program that represses B cell lymphoma development. Nature medicine, 21(10), 1199-1208.
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6

Cytokine-Mediated Signaling Pathway Inhibition

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RPMI-1640 medium was purchased from Invitrogen Life Technologies, CA, USA. Fetal bovine serum (FBS) was supplied by HyClone, UT, USA. Tyrphostin AG490 (a JAK2 kinase inhibitor), dimethyl sulfoxide (DMSO), and acetic acid were purchased from Sigma (Poole, UK). IL-6 was purchased from Pepro Tech (NJ, USA). The primary antibodies against total STAT3, COX-2, and β-actin and streptavidin/peroxidase for immunochemical staining were purchased from BIOS China, and the antibody against phosphorylated tyrosine705 STAT3 (p-tyr-705 STAT3) was purchased from Cell Signaling Technology (Massachusetts, USA). The anti-C/EBPβ antibody was purchased from Santa Cruz (California, USA). Cell culture dishes were purchased from NECU (Denmark). IL-6 was dissolved in acetic acid to a stock concentration of 1 μg/ml, and AG490 was dissolved in DMSO to a stock concentration of 100 mM/L. Both stock solutions were stored at −20°C for further use. For all experiments, the optimal working concentrations of the tested reagents were prepared by diluting with RPMI-1640 medium.
Ren M., McGowan E., Li Y., Zhu X., Lu X., Zhu Z., Lin Y, & He S. (2019). Saikosaponin-d Suppresses COX2 Through p-STAT3/C/EBPβ Signaling Pathway in Liver Cancer: A Novel Mechanism of Action. Frontiers in Pharmacology, 10, 623.
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7

Signaling Pathway Antibody Panel for JAK-STAT Analysis

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Antibodies against p-Tyr1022/1023-JAK1 (#3331), p-Tyr1007/1008-JAK2 (#3776), p-Tyr980/981-JAK3 (#5031), p-Tyr1054/1055-TYK2 (#9321), p-Tyr701-STAT1 (#9167), p-Tyr705-STAT3 (#9145), p-Tyr694-STAT5 (#9359), JAK1 (#3332S), JAK2 (#3230), JAK3 (#8863), TYK2 (#9312), STAT1 (#14994), STAT3 (#12640), STAT5 (#25656), c-Myc (#13987), cyclin D1 (#AF0931), Bcl-xL (#2764), p-Ser536-NF-κB (#3033), p-Thr308-AKT (#9275), p-Ser9-GSK-3β (#5558), p-Thr183/Tyr185-SAPK/JNK (#4668), NF-κB (#8242), AKT (#4691), GSK-3β (#9832), and SAPK/JNK (#9252) were purchased from Cell Signaling Technology (Danvers, MA, USA). Antibodies against α-tubulin were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA). Protease inhibitor (B14001) and phosphatase inhibitor (B15001) were purchased from Bimake (Houston, TX, USA). Recombinant human STAT3 protein (ab268982) was obtained from Abcam (Cambridge, Britain). Recombinant human interleukin-6 (IL-6) protein (200-06) was purchased from PeproTech (Rocky Hill, CT, USA). Nitrocellulose membranes and chemiluminescent horseradish peroxidase (HRP) substrate were obtained from Millipore (Billerica, MA, USA). Gefitinib was purchased from Selleckchem (Houston, TX, USA). DAB color solution, hematoxylin solution, and eosin staining solution were purchased from Servicebio (Wuhan, China).
He N., Li L., Li R., Zhang S.Q., Wu L.H., Guan X., Zhang Q.Y., Jiang T, & Yang J.B. (2023). A Novel Ageladine A Derivative Acts as a STAT3 Inhibitor and Exhibits Potential Antitumor Effects. International Journal of Molecular Sciences, 24(10), 8859.
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8

Western Blot Analysis of Apoptosis

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Cells were collected by centrifugation at 500 g for 5 min, and the pellets were resuspended in a lysis buffer (1% NP40, 1 mM phenylmethylsulfonyl fluoride, 40 mM Tris-HCl [pH 8.0], 150 mM NaCl, 1 mM NaOV) at 4°C for 15 min. Cell lysates (20 μg protein per lane) were fractionated on 12.5% SDS-polyacrylamide gels before being transferred to the membrane (Immobilon-P membranes [Merck Millipore, Billerica, MA, USA]) according to the standard protocol. Antibody binding was detected by using the enhanced chemiluminescence kit with hyper-ECL film (GE Healthcare Japan, Hino, Japan). Antibodies against caspase-3, carpase-8 and carpase-9, PARP, Bid, STAT3, pTyr705-STAT3, pTyr1007/1008-JAK2, Akt, p44/42 MAPK (Erk1/2) and NF-κB p65 were purchased from Cell Signaling Technology (Beverly, MA, USA), while those against Bcl-2, Bcl-xL, Mcl-1, RelB, c-Rel and β-actin were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA).
Sagawa M., Tabayashi T., Kimura Y., Tomikawa T., Nemoto-Anan T., Watanabe R., Tokuhira M., Ri M., Hashimoto Y., Iida S, & Kizaki M. (2015). TM-233, a novel analog of 1′-acetoxychavicol acetate, induces cell death in myeloma cells by inhibiting both JAK/STAT and proteasome activities. Cancer Science, 106(4), 438-446.
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9

Immunohistochemistry Protocol for HCC Tissue Microarray

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Tissue microarray (TMA) containing matched pairs of tumor and peritumoral liver tissues from 13 patients with primary HCC was constructed by Shanghai Biochip Co., Ltd, as previously described.45 (link) Briefly, the sections were dewaxed with xylene and dehydrated in a graded series of ethanol. After neutralization of endogenous peroxidase and microwave antigen retrieval, the sections were blocked with goat serum and incubated overnight in primary antibody against COX-2 (#12282, Cell Signaling Technology), p-Tyr705-STAT3 (#9145, Cell Signaling Technology) or TLR4 (sc-10741, Santa Cruz Biotechnology). After washing by PBS containing 0.05% Tween 20, the sections were incubated with second antibody for 30 min at room temperature, and then treated with HRP-conjugated streptavidin. The reaction products were visualized using 3-3-diamino-benzidine-tetrahydrochloride (DAB) followed by nuclei staining in hematoxylin. The stained area and Integrated Optical Density (IOD) of four random fields in each section were calculated using Image-pro Plus 6.0 software. The density mean, equal to (IOD SUM)/area, represented the exact protein expression.
Lin A., Wang G., Zhao H., Zhang Y., Han Q., Zhang C., Tian Z, & Zhang J. (2015). TLR4 signaling promotes a COX-2/PGE2/STAT3 positive feedback loop in hepatocellular carcinoma (HCC) cells. Oncoimmunology, 5(2), e1074376.
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10

Arginase1 and STAT3 Phosphorylation

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Arginase1 antibody was obtained from Thermofisher (Cat # 711765). Cells were treated with MTA for 24 h and cells were lysed with RIPA buffer and ran on a Bio-Rad ready-made gel and transferred to nitrocellulose membrane. Total STAT3 and p-Tyr705 STAT3 were obtained from Cell Signaling Technologies (Cat # 9139, 9145).
Hansen L.J., Yang R., Roso K., Wang W., Chen L., Yang Q., Pirozzi C.J, & He Y. (2022). MTAP loss correlates with an immunosuppressive profile in GBM and its substrate MTA stimulates alternative macrophage polarization. Scientific Reports, 12, 4183.
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