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      PE-CF594 Mouse Anti-Stat5 (pY694)
      PE-CF594 Mouse Anti-Stat5 (pY694)
      Flow cytometric analysis of Stat5 (pY694) expression in IL-2-treated human peripheral blood lymphocytes. Human whole blood was either stimulated with 100 ng/ml (final concentration) of BD Pharmingen™ Recombinant Human IL-2 (Cat. No. 554903) for 15 minutes at 37ºC (solid line histogram) or was left unstimulated (dashed line histogram). The erythrocytes were lysed and the leukocytes were fixed with 1× BD Phosflow™ Lyse/Fix Buffer (Cat. No. 558049) for 15 minutes at 37ºC. The leukocytes were washed, permeabilized (BD Phosflow™ Perm Buffer III, Cat. No. 558050) on ice for 30 minutes, washed and then stained with BD Phosflow™ PE-CF594 anti-Stat5 (pY694) (Cat. No.562501). The fluorescence histograms were derived from events with the forward and side light-scatter characteristics of intact lymphocytes. Flow cytometry was performed using a BD™ LSR II Flow Cytometer System.
      PE-CF594 Mouse Anti-Stat5 (pY694)
      Flow cytometric analysis of Stat5 (pY694) expression in IL-2-treated human peripheral blood lymphocytes. Human whole blood was either stimulated with 100 ng/ml (final concentration) of BD Pharmingen™ Recombinant Human IL-2 (Cat. No. 554903) for 15 minutes at 37ºC (solid line histogram) or was left unstimulated (dashed line histogram). The erythrocytes were lysed and the leukocytes were fixed with 1× BD Phosflow™ Lyse/Fix Buffer (Cat. No. 558049) for 15 minutes at 37ºC. The leukocytes were washed, permeabilized (BD Phosflow™ Perm Buffer III, Cat. No. 558050) on ice for 30 minutes, washed and then stained with BD Phosflow™ PE-CF594 anti-Stat5 (pY694) (Cat. No.562501). The fluorescence histograms were derived from events with the forward and side light-scatter characteristics of intact lymphocytes. Flow cytometry was performed using a BD™ LSR II Flow Cytometer System.
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      BD Phosflow™
      Signal transducer and activator of transcription 5; MGF; MPF
      Human (QC Testing)
      Mouse IgG1, κ
      Phosphorylated Human Phosphorylated Stat5 Peptide
      Intracellular staining (flow cytometry) (Routinely Tested)
      5 µl
      AB_2737628
      Aqueous buffered solution containing BSA and ≤0.09% sodium azide.
      RUO


      Preparation and Storage

      Store undiluted at 4°C and protected from prolonged exposure to light. Do not freeze. The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography. The antibody was conjugated with BD Horizon™ PE-CF594 under optimum conditions, and unconjugated antibody and free PE-CF594 were removed.
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      推奨アッセイ手順

      This antibody conjugate is suitable for intracellular staining of human whole blood (using BD Phosflow™ Lyse/Fix Buffer) and peripheral blood mononuclear cells (using BD Cytofix™ Fixation Buffer or BD Phosflow™ Fix Buffer I).

      This purified or conjugated mAb was characterized by flow cytometry (Flow) and western blot analysis (WB) using these model systems:

      Method                        Species                Cells                        Treatment        Fixation                                Perm buffer                Result

      Flow                        Human                PBMC                        IL-2                        Fixation Buffer                III                                Positive Staining

      Flow                        Human                PBMC                        IL-2                        Fixation Buffer                I or II                        Unsatisfactory

      Flow                        Human                Whole Blood                IL-2                        Lyse/Fix                                III                                Positive Staining

      Flow                        Human                Whole Blood                IL-2                        Lyse/Fix                                 I or II                        Unsatisfactory

      WB                                Human                A431 Cell Lysate        EGF                        Not Applicable                Not Applicable        92 kDa

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      Product Notices

      1. This reagent has been pre-diluted for use at the recommended Volume per Test. We typically use 1 × 10^6 cells in a 100-µl experimental sample (a test).
      2. Source of all serum proteins is from USDA inspected abattoirs located in the United States.
      3. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
      4. Please observe the following precautions: Absorption of visible light can significantly alter the energy transfer occurring in any tandem fluorochrome conjugate; therefore, we recommend that special precautions be taken (such as wrapping vials, tubes, or racks in aluminum foil) to prevent exposure of conjugated reagents, including cells stained with those reagents, to room illumination.
      5. Caution: Sodium azide yields highly toxic hydrazoic acid under acidic conditions. Dilute azide compounds in running water before discarding to avoid accumulation of potentially explosive deposits in plumbing.
      6. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
      7. Texas Red is a registered trademark of Molecular Probes, Inc., Eugene, OR.
      8. CF™ is a trademark of Biotium, Inc.
      9. When excited by the yellow-green (561-nm) laser, the fluorescence may be brighter than when excited by the blue (488-nm) laser.
      10. This product is provided under an Agreement between BIOTIUM and BD Biosciences. The manufacture, use, sale, offer for sale, or import of this product is subject to one or more patents or pending applications owned or licensed by Biotium, Inc. This product, and only in the amount purchased by buyer, may be used solely for buyer’s own internal research, in a manner consistent with the accompanying product literature. No other right to use, sell or otherwise transfer (a) this product, or (b) its components is hereby granted expressly, by implication or by estoppel. This product is for research use only. Diagnostic uses require a separate license from Biotium, Inc. For information on purchasing a license to this product including for purposes other than research, contact Biotium, Inc., 3159 Corporate Place, Hayward, CA 94545, Tel: (510) 265-1027. Fax: (510) 265-1352. Email: btinfo@biotium.com.
      11. Because of the broad absorption spectrum of the tandem fluorochrome, extra care must be taken when using multi-laser cytometers, which may directly excite both PE and CF™594.
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      562501 Rev. 1
      抗体の詳細
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      47/Stat5(pY694)

      Stat (Signal transducer and activators of transcription) proteins mediate the biological activity of cytokines, including interleukins, interferons, erythropoietin, and growth factors. Ligand-receptor interaction activates constitutively-associated JAK family kinases and subsequent recruitment/activation of Stat proteins by tyrosine phosphorylation. Active Stat proteins then move to the nucleus to promote transcription of cytokine-inducible genes. Seven Stat proteins have been cloned, each of which is differentially expressed and/or activated in a cytokine-specific and cell type-specific manner. Stat5 has been characterized and shown to be encoded by two separate genes, Stat5a and Stat5b, which share over 90% identity at the amino acid level. Stat5a has been shown to be involved in lactogenesis and mammary development, while Stat5b has been shown to be involved in growth hormone signaling and liver gene expression. Both Stat5a and Stat5b are involved in IL-2 induced peripheral T cell proliferation. The peptide hormone, prolactin, binds to the prolactin receptor (PRLR) to initiate the lactogenic response. There are at least three forms of PRLR; however, only the long form activates the 92-kDa Stat5 protein by inducing phosphorylation at Y694. Once phosphorylated, Stat5 becomes an essential transcription factor which binds to the β-casein gene promoter. The presence of an SH2 domain within Stat5 suggests that it may directly interact with protein tyrosine kinases (PTKs) such as JAK2.

      The 47 monoclonal antibody recognizes the phosphorylated Y694 of Stat5a. The homologous phosphorylation site in Stat5b is Y699.

      This antibody is conjugated to BD Horizon™ PE-CF594, which has been developed exclusively by BD Biosciences as a better alternative to PE-Texas Red®. PE-CF594 excites and emits at similar wavelengths to PE-Texas Red® yet exhibits improved brightness and spectral characteristics. Due to PE having maximal absorption peaks at 496 nm and 564 nm, PE-CF594 can be excited by the blue (488-nm), green (532-nm) and yellow-green (561-nm) lasers and can be detected with the same filter set as PE-Texas Red® (eg 610/20-nm filter).

      562501 Rev. 1
      フォーマットの詳細
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      PE-CF594
      BD Horizon™ PE-CF594 dye is a part of the BD PE family of dyes. This tandem fluorochrome is comprised of a R-Phycoerythrin (PE) donor that has excitation maxima (Ex Max) of 496-nm and 566-nm and an acceptor dye with an emission maximum (Em Max) at 615-nm. PE-CF594, driven by BD innovation, is designed to be excited by the blue (488-nm), Green (532-nm) and yellow-green (561-nm) lasers and detected using an optical filter centered near 615 nm (e.g., a 610/20-nm bandpass filter). The donor dye can be excited by the Blue (488-nm), Green (532-nm) and yellow-green (561-nm) lasers and the acceptor dye can be excited by the green (532-nm) laser resulting in cross-laser excitation and fluorescence spillover. Please ensure that your instrument’s configurations (lasers and optical filters) are appropriate for this dye.
      altImg
      PE-CF594
      Yellow-Green 488 nm, 532 nm, 561 nm
      496 nm, 566 nm
      615 nm
      562501 Rev.1
      引用＆参考文献
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      Development References (5)

      1. Bromberg J, Darnell JE. The role of STATs in transcriptional control and their impact on cellular function. Oncogene. 2000; 19(21):2468-2473. (Biology). View Reference
      2. Gouilleux F, Wakao H, Mundt M, Groner B. Prolactin induces phosphorylation of Tyr694 of Stat5 (MGF), a prerequisite for DNA binding and induction of transcription. EMBO J. 1994; 13(18):4361-4369. (Biology). View Reference
      3. Liu KD, Gaffen SL, Goldsmith MA. JAK/STAT signaling by cytokine receptors. Curr Opin Immunol. 1998; 10(3):271-278. (Biology). View Reference
      4. Van De Wiele CJ, Marino JH, Murray BW, Vo SS, Whetsell ME, Teague TK. Thymocytes between the -Selection and Positive Selection Checkpoints Are Nonresponsive to IL-7 as Assessed by STAT-5 Phosphorylation. J Immunol. 2004; 172(7):4235-4244. (Biology). View Reference
      5. Wakao H, Gouilleux F, Groner B. Mammary gland factor (MGF) is a novel member of the cytokine regulated transcription factor gene family and confers the prolactin response. EMBO J. 1994; 13(9):2182-2191. (Biology). View Reference
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      562501 Rev. 1

      Please refer to Support Documents for Quality Certificates


      Global - Refer to manufacturer's instructions for use and related User Manuals and Technical data sheets before using this products as described


      Comparisons, where applicable, are made against older BD Technology, manual methods or are general performance claims.  Comparisons are not made against non-BD technologies, unless otherwise noted.

      For Research Use Only. Not for use in diagnostic or therapeutic procedures.

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