Antibody data
- Antibody Data
- Antigen structure
- References [17]
- Comments [0]
- Validations
- Flow cytometry [1]
- Other assay [6]
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- Product number
- 12-9010-42 - Provider product page
- Provider
- Invitrogen Antibodies
- Product name
- Phospho-STAT5 (Tyr694) Monoclonal Antibody (SRBCZX), PE, eBioscience™
- Antibody type
- Monoclonal
- Antigen
- Other
- Description
- Description: This SRBCZX monoclonal antibody recognizes signal transducer and activator of transcription 5 (STAT5) when phosphorylated on tyrosine 694. STAT proteins are activated by ligand binding to receptors, such as cytokine receptors, that associate with Janus kinase (JAK) family members. Following their phosphorylation by JAKs, STAT proteins translocate to the nucleus where they bind to DNA and regulate transcription of specific genes in a cell type- and cytokine-specific manner. In response to cytokines that signal through the common gamma chain such as IL-2, IL-7, and IL-15, STAT5 is phosphorylated on tyrosine 694 by JAK1 and JAK3. Cytokines such as IL-3, IL-5, and GM-CSF that signal via the common beta chain induce STAT5 phosphorylation on tyrosine 694 by JAK 2. Phosphorylation of STAT5 on tyrosine 694 is essential for STAT5 dimer formation, nuclear translocation, and DNA binding activity. Specificity of this SRBCZX clone was determined by ELISA and flow cytometry. Applications Reported: This SRBCZX antibody has been reported for use in intracellular staining followed by flow cytometric analysis. Applications Tested: This SRBCZX antibody has been pre-titrated and tested by intracellular staining followed by flow cytometric analysis of stimulated normal human peripheral blood cells. This can be used at 5 µL (0.25 µg) per test. A test is defined as the amount (µg) of antibody that will stain a cell sample in a final volume of 100 µL. Cell number should be determined empirically but can range from 10^5 to 10^8 cells/test. Staining Protocol: We recommend using Protocol C: Two-step protocol: Fixation/Methanol. Protocol A: Two-step protocol: intracellular (cytoplasmic) proteins and Protocol B: One-step protocol: intracellular (nuclear) proteins cannot be used. All Protocols can be found in the Flow Cytometry Protocols: "Staining Intracellular Antigens for Flow Cytometry Protocol" located in the Best Protocols Section under the Resources tab online. Excitation: 488-561 nm; Emission: 578 nm; Laser: Blue Laser, Green Laser, Yellow-Green Laser. Filtration: 0.2 µm post-manufacturing filtered.
- Reactivity
- Human, Mouse
- Host
- Mouse
- Conjugate
- Yellow dye
- Isotype
- IgG
- Antibody clone number
- SRBCZX
- Vial size
- 100 Tests
- Concentration
- 5 µL/Test
- Storage
- 4° C, store in dark, DO NOT FREEZE!
Submitted references GM-CSF impairs erythropoiesis by disrupting erythroblastic island formation via macrophages.
Interleukin-23 receptor defines T helper 1-like regulatory T cells in oral squamous cell carcinoma.
Aged neutrophils form mitochondria-dependent vital NETs to promote breast cancer lung metastasis.
Ageing promotes early T follicular helper cell differentiation by modulating expression of RBPJ.
SRC-3 Functions as a Coactivator of T-bet by Regulating the Maturation and Antitumor Activity of Natural Killer Cells.
Novel cytokine-antibody fusion protein, N-820, to enhance the functions of ex vivo expanded natural killer cells against Burkitt lymphoma.
Long-Term Programming of CD8 T Cell Immunity by Perinatal Exposure to Glucocorticoids.
ILC3 deficiency and generalized ILC abnormalities in DOCK8-deficient patients.
Receptor-mediated dimerization of JAK2 FERM domains is required for JAK2 activation.
CISH promoter polymorphism effects on T cell cytokine receptor signaling and type 1 diabetes susceptibility.
Erythropoietin enhances Kupffer cell number and activity in the challenged liver.
Aberrant plasma IL-7 and soluble IL-7 receptor levels indicate impaired T-cell response to IL-7 in human tuberculosis.
Expression and function of hematopoiesis-stimulating factor receptors on the GPI(-) and GPI(+) hematopoietic stem cells of patients with paroxysmal nocturnal hemoglobinuria/aplastic anemia syndrome.
Monomethylarsonous acid (MMA+3) Inhibits IL-7 Signaling in Mouse Pre-B Cells.
Murine iPSC-Derived Macrophages as a Tool for Disease Modeling of Hereditary Pulmonary Alveolar Proteinosis due to Csf2rb Deficiency.
Environmentally relevant concentrations of arsenite and monomethylarsonous acid inhibit IL-7/STAT5 cytokine signaling pathways in mouse CD3+CD4-CD8- double negative thymus cells.
Combination of imatinib with CXCR4 antagonist BKT140 overcomes the protective effect of stroma and targets CML in vitro and in vivo.
Cao W, Fan W, Wang F, Zhang Y, Wu G, Shi X, Shi JX, Gao F, Yan M, Guo R, Li Y, Li W, Du C, Jiang Z
Journal of translational medicine 2022 Jan 3;20(1):11
Journal of translational medicine 2022 Jan 3;20(1):11
Interleukin-23 receptor defines T helper 1-like regulatory T cells in oral squamous cell carcinoma.
Li W, An N, Wang M, Liu X, Mei Z
Immunity, inflammation and disease 2022 Dec;10(12):e746
Immunity, inflammation and disease 2022 Dec;10(12):e746
Aged neutrophils form mitochondria-dependent vital NETs to promote breast cancer lung metastasis.
Yang C, Wang Z, Li L, Zhang Z, Jin X, Wu P, Sun S, Pan J, Su K, Jia F, Zhang L, Wang H, Yu X, Shao X, Wang K, Qiu F, Yan J, Huang J
Journal for immunotherapy of cancer 2021 Oct;9(10)
Journal for immunotherapy of cancer 2021 Oct;9(10)
Ageing promotes early T follicular helper cell differentiation by modulating expression of RBPJ.
Webb LMC, Fra-Bido S, Innocentin S, Matheson LS, Attaf N, Bignon A, Novarino J, Fazilleau N, Linterman MA
Aging cell 2021 Jan;20(1):e13295
Aging cell 2021 Jan;20(1):e13295
SRC-3 Functions as a Coactivator of T-bet by Regulating the Maturation and Antitumor Activity of Natural Killer Cells.
Hu M, Lu Y, Qi Y, Zhang Z, Wang S, Xu Y, Chen F, Tang Y, Chen S, Chen M, Du C, Shen M, Wang F, Su Y, Deng Y, Wang J
Cancer immunology research 2020 Sep;8(9):1150-1162
Cancer immunology research 2020 Sep;8(9):1150-1162
Novel cytokine-antibody fusion protein, N-820, to enhance the functions of ex vivo expanded natural killer cells against Burkitt lymphoma.
Chu Y, Nayyar G, Kham Su N, Rosenblum JM, Soon-Shiong P, Lee J, Safrit JT, Barth M, Lee D, Cairo MS
Journal for immunotherapy of cancer 2020 Oct;8(2)
Journal for immunotherapy of cancer 2020 Oct;8(2)
Long-Term Programming of CD8 T Cell Immunity by Perinatal Exposure to Glucocorticoids.
Hong JY, Lim J, Carvalho F, Cho JY, Vaidyanathan B, Yu S, Annicelli C, Ip WKE, Medzhitov R
Cell 2020 Mar 5;180(5):847-861.e15
Cell 2020 Mar 5;180(5):847-861.e15
ILC3 deficiency and generalized ILC abnormalities in DOCK8-deficient patients.
Eken A, Cansever M, Okus FZ, Erdem S, Nain E, Azizoglu ZB, Haliloglu Y, Karakukcu M, Ozcan A, Devecioglu O, Aksu G, Arikan Ayyildiz Z, Topal E, Karakoc Aydiner E, Kiykim A, Metin A, Cipe F, Kaya A, Artac H, Reisli I, Guner SN, Uygun V, Karasu G, Dönmez Altuntas H, Canatan H, Oukka M, Ozen A, Chatila TA, Keles S, Baris S, Unal E, Patiroglu T
Allergy 2020 Apr;75(4):921-932
Allergy 2020 Apr;75(4):921-932
Receptor-mediated dimerization of JAK2 FERM domains is required for JAK2 activation.
Ferrao RD, Wallweber HJ, Lupardus PJ
eLife 2018 Jul 25;7
eLife 2018 Jul 25;7
CISH promoter polymorphism effects on T cell cytokine receptor signaling and type 1 diabetes susceptibility.
Seyfarth J, Ahlert H, Rosenbauer J, Baechle C, Roden M, Holl RW, Mayatepek E, Meissner T, Jacobsen M
Molecular and cellular pediatrics 2018 Feb 6;5(1):2
Molecular and cellular pediatrics 2018 Feb 6;5(1):2
Erythropoietin enhances Kupffer cell number and activity in the challenged liver.
Gilboa D, Haim-Ohana Y, Deshet-Unger N, Ben-Califa N, Hiram-Bab S, Reuveni D, Zigmond E, Gassmann M, Gabet Y, Varol C, Neumann D
Scientific reports 2017 Sep 4;7(1):10379
Scientific reports 2017 Sep 4;7(1):10379
Aberrant plasma IL-7 and soluble IL-7 receptor levels indicate impaired T-cell response to IL-7 in human tuberculosis.
Lundtoft C, Afum-Adjei Awuah A, Rimpler J, Harling K, Nausch N, Kohns M, Adankwah E, Lang F, Olbrich L, Mayatepek E, Owusu-Dabo E, Jacobsen M
PLoS pathogens 2017 Jun;13(6):e1006425
PLoS pathogens 2017 Jun;13(6):e1006425
Expression and function of hematopoiesis-stimulating factor receptors on the GPI(-) and GPI(+) hematopoietic stem cells of patients with paroxysmal nocturnal hemoglobinuria/aplastic anemia syndrome.
Fu R, Ding SX, Liu YI, Li LJ, Liu H, Wang HL, Zhang T, Shao ZH
Experimental and therapeutic medicine 2016 May;11(5):1668-1672
Experimental and therapeutic medicine 2016 May;11(5):1668-1672
Monomethylarsonous acid (MMA+3) Inhibits IL-7 Signaling in Mouse Pre-B Cells.
Ezeh PC, Xu H, Lauer FT, Liu KJ, Hudson LG, Burchiel SW
Toxicological sciences : an official journal of the Society of Toxicology 2016 Feb;149(2):289-99
Toxicological sciences : an official journal of the Society of Toxicology 2016 Feb;149(2):289-99
Murine iPSC-Derived Macrophages as a Tool for Disease Modeling of Hereditary Pulmonary Alveolar Proteinosis due to Csf2rb Deficiency.
Mucci A, Kunkiel J, Suzuki T, Brennig S, Glage S, Kühnel MP, Ackermann M, Happle C, Kuhn A, Schambach A, Trapnell BC, Hansen G, Moritz T, Lachmann N
Stem cell reports 2016 Aug 9;7(2):292-305
Stem cell reports 2016 Aug 9;7(2):292-305
Environmentally relevant concentrations of arsenite and monomethylarsonous acid inhibit IL-7/STAT5 cytokine signaling pathways in mouse CD3+CD4-CD8- double negative thymus cells.
Xu H, Lauer FT, Liu KJ, Hudson LG, Burchiel SW
Toxicology letters 2016 Apr 15;247:62-8
Toxicology letters 2016 Apr 15;247:62-8
Combination of imatinib with CXCR4 antagonist BKT140 overcomes the protective effect of stroma and targets CML in vitro and in vivo.
Beider K, Darash-Yahana M, Blaier O, Koren-Michowitz M, Abraham M, Wald H, Wald O, Galun E, Eizenberg O, Peled A, Nagler A
Molecular cancer therapeutics 2014 May;13(5):1155-69
Molecular cancer therapeutics 2014 May;13(5):1155-69
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Supportive validation
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- Invitrogen Antibodies (provider)
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- Experimental details
- LEFT: Intracellular staining of untreated (orange histogram) or 15-minute IL-2-treated (purple histogram) human Th2-polarized CD4+ with Anti-Human/Mouse phospho-STAT5 (Y694) PE. Cells in the lymphocyte gate were used for analysis. RIGHT: Intracellular staining of untreated (orange histogram) or 15-minute GM-CSF-treated (purple histogram) mouse thioglycolate-elicited peritoneal exudate cells with Anti-Human/Mouse phospho-STAT5 (Y694) PE. CD11c+ cells in the large scatter population were used for analysis. In both panels, cells were stained using the Fixation/Methanol protocol.
- Conjugate
- Yellow dye
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- NULL
- Conjugate
- Yellow dye
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Figure 1 EPO regulates EPO-R expression and elicits downstream signalling in RKC-2 cells. All graphs represent mean +- SEM. ( A-B) RKC-2 cells were cultured in the presence or absence of 5 U/ml EPO for 24 h. ( A ) EPO-R transcript levels were evaluated by RT-PCR, N = 5-8, *p < 0.05. Rat BMDM (positive control) were considered as 1. ( B ) EPO-R surface expression was evaluated by flow cytometry. Top: Grey and black line histograms depict surface EPO-R in control (considered as 1) and EPO-treated cells, respectively. Full histogram depicts FITC conjugated Goat anti mouse antibody. Bottom: Quantification of surface EPO-R, N = 7, **p < 0.01. ( C-D ) RKC-2 cells were stimulated for 0, 10 and 30 min with 10 U/ml EPO. Top: Grey, black and dashed histograms represent 0, 10 and 30 min EPO-treated cells, respectively. Bottom: The graphs depict quantification of mean fluorescence intensity (MFI) of flow cytometry analysis for p-ERK1/2 and p-STAT5, ( C ) and ( D ), respectively. Levels at 0 min were considered as 1, N = 4-5, **p < 0.01, *p < 0.05.
- Conjugate
- Yellow dye
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Figure 5. EPOR and LEPR switch residues are required for Epo and Leptin-induced STAT phosphorylation. ( a,b ) Stable Ba/F3 cell lines expressing wild-type, full length mouse EPOR or LEPR were generated and analyzed for STAT phosphorylation by flow cytometry after stimulation with EPO or leptin. ( a ) Representative plots comparing phospho-STAT5 staining of parental Ba/F3 cells or Ba/F3 cells expressing EPOR. Cells were stimulated with 1 nM mouse EPO for 15 min before fixation, staining, and analysis. ( b ) Representative plots comparing phospho-STAT3 staining of parental Ba/F3 cells or Ba/F3 cells expressing LEPR. Cells were stimulated with 100 pM mouse Leptin for 4 hr prior to fixation, staining, and analysis. ( c ) Analysis of STAT5 phosphorylation by flow cytometry for EPOR wild-type, switch region, and box1 mutants, stimulated with 1 nM EPO as in ( a ). Mean levels of STAT5 phosphorylation were assessed in three separate experiments, with wild-type signal representing 100% in all three experiments. ( d ) Analysis of STAT3 phosphorylation by flow cytometry for LEPR wild-type, switch region, and box1 mutants, stimulated with 100 pM Leptin, as in ( b ). Mean levels of STAT3 phosphorylation was assessed in three separate experiments, with wild-type signal representing 100% in all three experiments. Error bars represent standard error of the mean (SEM). Figure 5--figure supplement 1. Analysis of Ba/F3 stable cell lines expressing EPOR or LEPR mutants. ( a ) Alignment of human
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- Yellow dye
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Figure 5--figure supplement 1. Analysis of Ba/F3 stable cell lines expressing EPOR or LEPR mutants. ( a ) Alignment of human and mouse sequences for EPOR and LEPR. Note the numbering differences for mouse versus human (minus one residue for EPOR and minus two residues for LEPR). Residues in the switch region and box1, which are mutated in our experiments, are completely conserved between human and mouse. ( b-c ) Receptor surface staining for stable cell lines expressing ( b ) EPOR variants and ( c ) LEPR variants. Plots represent cumulative events (Y axis) at or below a given staining level (X axis). ( d ) Representative raw flow cytometry histograms showing the phospho-STAT5 (EPOR) or phospho-STAT3 (LEPR) staining in unstimulated cells (black trace) or cells stimulated with cytokine (red trace) as described in Materials and methods.
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- Yellow dye
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- Invitrogen Antibodies (provider)
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- Figure 3. The P-STAT5 MFI in the (A) GPI + cells and GPI - cells of PNH/AA patients and (B) GPI + cells of normal controls with or without stimulation by G-CSF or SCF. (C) The MFI of P-STAT5 in GPI + and GPI - cells of the bone marrow before and after G-CSF or SCF stimulation. G-CSF, granulocyte colony-stimulating factor; SCF, stem cell factor; P-STAT5, phosphorylated signal transducer and activator of transcription 5; MFI, mean fluorescence intensity; GPI, glycophosphatidylinositol; PNH, paroxysmal nocturnal hemoglobinuria; CD, cluster of differentiation.
- Conjugate
- Yellow dye
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Figure 6 Disease Modeling of herPAP Using miPAP iPSCs (A) Number of CD41 + progenitor-derived colonies in methylcellulose-based clonogenic assays employing complete (IL-6, erythropoietin, SCF, IL-3, and supplemented with 20 ng/ml human G-CSF + 20 ng/ml murine GM-CSF) or basic medium (50 ng/ml murine GM-CSF only; independent experiments, n = 3 miPAP1 and 2, n = 2 CD45.1 iPSC, mean +- SD) and representative pictures of colonies. Scale bars, 500 mum. (B) GM-CSF clearance assay comparing miPAP-Mphi with CD45.1(10.3) iPSC-Mphi, BMlin - -Mphi, and no cells incubated with 2 ng/ml GM-CSF: at indicated time points (0, 4, 10, 24, and 30 hr) GM-CSF concentrations in supernatants were analyzed by ELISA, normalized to 0 hr (independent experiments, n = 3 BMlin - and CD45.1 iPSC, n = 2 miPAP1 and 2, mean +- SD). (C and D) Flow cytometry plots of STAT5 phosphorylation levels upon stimulation with mGM-CSF (C) and (D) summary of mean fluorescence intensity (MFI) data (independent experiments, n = 3 BMlin - and CD45.1 iPSC, n = 2 miPAP1 and 2, mean +- SD). ns, not significant; ** p
- Conjugate
- Yellow dye