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PE-CF594 Mouse Anti-Stat3 (pY705)
PE-CF594 Mouse Anti-Stat3 (pY705)
Analysis of Stat3 (pY705) expressed in human peripheral blood lymphocytes. Whole blood was either left unstimulated (dashed line histogram) or stimulated (solid line histogram) with 100 ng/ml BD Pharmingen™ Recombinant Human IL-6 (Cat. No. 550071) for 15 minutes at 37°C. Cells were lysed and fixed in a single step using BD Phosflow™ Lyse/Fix Buffer (Cat. No. 558049) for 10 min at 37 °C. Cells were then permeabilized in BD Phosflow™ Perm Buffer III (Cat. No. 558050) on ice for 30 minutes and then stained with BD Horizon™ PE-CF594 anti-Stat3 (pY705) mAb (Cat. No. 562673). The fluorescence histograms were derived from gated events with the forward and side light-scatter characteristics of intact lymphocytes. Flow cytometry was performed using a BD™ LSR II Flow Cytometer System.
Analysis of Stat3 (pY705) expressed in human peripheral blood lymphocytes. Whole blood was either left unstimulated (dashed line histogram) or stimulated (solid line histogram) with 100 ng/ml BD Pharmingen™ Recombinant Human IL-6 (Cat. No. 550071) for 15 minutes at 37°C. Cells were lysed and fixed in a single step using BD Phosflow™ Lyse/Fix Buffer (Cat. No. 558049) for 10 min at 37 °C. Cells were then permeabilized in BD Phosflow™ Perm Buffer III (Cat. No. 558050) on ice for 30 minutes and then stained with BD Horizon™ PE-CF594 anti-Stat3 (pY705) mAb (Cat. No. 562673). The fluorescence histograms were derived from gated events with the forward and side light-scatter characteristics of intact lymphocytes. Flow cytometry was performed using a BD™ LSR II Flow Cytometer System.
Product Details
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BD Phosflow™
Acute-phase response factor, APRF
Human (QC Testing), Mouse (Tested in Development)
Mouse IgG2a, κ
Phosphorylated Human Stat3 Peptide
Intracellular staining (flow cytometry) (Routinely Tested)
5 µl
AB_2737714
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.

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. An isotype control should be used at the same concentration as the antibody of interest.
  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.
  12. Species cross-reactivity detected in product development may not have been confirmed on every format and/or application.
  13. Source of all serum proteins is from USDA inspected abattoirs located in the United States.
562673 Rev. 1
Antibody Details
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4/P-STAT3

Stat (Signal transducer and activators of transcription) proteins are critical mediators of the biologic activity of cytokines, including interleukins, interferons, erythropoietin, and growth factors.  Ligand-receptor interaction leads to activation of 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. Stat3 is a 92-kDa protein that is activated as a DNA- binding protein through cytokines, such as IL-6, and growth factors, such as EGF. Stat3 activation occurs via tyrosine phosphorylation at Y705. Tyrosine phosphorylation in response to cytokine stimulation is generally mediated by JAK1. Upon activation, Stat3 dimerizes, translocates to the nucleus and binds DNA response elements, thereby regulating gene expression. It has been reported that Stat3 binds to DNA as a homodimer, but it is also capable of binding as a heterodimer with Stat1. In addition to tyrosine phosphorylation, Stat3 is also phosphorylated at S727 via the MAPK pathway.  Stat3 is widely expressed and can bind to the sis-inducible element (SIE) site from the c-fos promoter. This site is similar to the GAS element that is present in IFN-γ induced genes. Thus, phosphorylation of Y705 in Stat3 occurs in response to growth factors and cytokines, and is essential for normal transcription activity.

The 4/P-STAT3 monoclonal antibody recognizes the phosphorylated Y705 of Stat3.

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).

562673 Rev. 1
Format Details
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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.
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PE-CF594
Yellow-Green 488 nm, 532 nm, 561 nm
496 nm, 566 nm
615 nm
562673 Rev.1
Citations & References
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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. Imada K, Leonard WJ. The Jak-STAT pathway. Mol Immunol. 2000; 37:1-11. (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. Renner ED, Rylaarsdam S, Anover-Sombke S, et al. Novel signal transducer and activator of transcription 3 (STAT3) mutations, reduced T(H)17 cell numbers, and variably defective STAT3 phosphorylation in hyper-IgE syndrome. J Allergy Clin Immunol. 2008; 122(1):181-187. (Clone-specific: Flow cytometry). View Reference
  5. Tanaka S, Saito Y, Kunisawa J, et al. Development of mature and functional human myeloid subsets in hematopoietic stem cell-engrafted NOD/SCID/IL2rgammaKO mice. J Immunol. 2012; 188(12):6145-6155. (Clone-specific: Flow cytometry). View Reference
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562673 Rev. 1

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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.