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      PerCP-Cy™5.5 Mouse Anti-Stat3
      PerCP-Cy™5.5 Mouse Anti-Stat3
      Flow cytometric analysis of Stat3 expression in human peripheral blood lymphocytes. Whole blood was treated with BD Phosflow™ Lyse/Fix Buffer (Cat. No. 558049) to lyse erythrocytes and fix leucocytes. The cells were subsequently permeabilized with BD Phosflow™ Perm Buffer III (Cat. No. 558050; on ice, 30 min). The cells were then washed with BD Pharmingen™ Stain Buffer (Cat. No. 554656/554657) and stained with either PerCP-Cy™5.5 Mouse IgG1, κ Isotype Control (Cat. No. 550795, dashed line histogram) or PerCP-Cy™5.5 Mouse Anti-Stat3 antibody (Cat. No. 564133; solid line histogram). The fluorescence histograms were derived from gated events with the forward and side light-scatter characteristics of intact lymphocytes. Flow cytometric analysis was performed using a BD™ LSR II Flow Cytometer System.
      PerCP-Cy™5.5 Mouse Anti-Stat3
      Flow cytometric analysis of Stat3 expression in human peripheral blood lymphocytes. Whole blood was treated with BD Phosflow™ Lyse/Fix Buffer (Cat. No. 558049) to lyse erythrocytes and fix leucocytes. The cells were subsequently permeabilized with BD Phosflow™ Perm Buffer III (Cat. No. 558050; on ice, 30 min). The cells were then washed with BD Pharmingen™ Stain Buffer (Cat. No. 554656/554657) and stained with either PerCP-Cy™5.5 Mouse IgG1, κ Isotype Control (Cat. No. 550795, dashed line histogram) or PerCP-Cy™5.5 Mouse Anti-Stat3 antibody (Cat. No. 564133; solid line histogram). The fluorescence histograms were derived from gated events with the forward and side light-scatter characteristics of intact lymphocytes. Flow cytometric analysis was performed using a BD™ LSR II Flow Cytometer System.
      Product Details
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      BD Phosflow™
      STAT3; Acute-phase response factor; APRF; HIES
      Human (QC Testing), Mouse, Rat (Predicted)
      Mouse BALB/c IgG1, λ
      Human Stat3 Recombinant Protein
      Intracellular staining (flow cytometry) (Routinely Tested)
      5 µl
      AB_2738614
      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 PerCP-Cy5.5 under optimum conditions, and unconjugated antibody and free PerCP-Cy5.5 were removed. Storage of PerCP-Cy5.5 conjugates in unoptimized diluent is not recommended and may result in loss of signal intensity.
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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. An isotype control should be used at the same concentration as the antibody of interest.
      3. Source of all serum proteins is from USDA inspected abattoirs located in the United States.
      4. 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.
      5. 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.
      6. PerCP-Cy5.5–labelled antibodies can be used with FITC- and R-PE–labelled reagents in single-laser flow cytometers with no significant spectral overlap of PerCP-Cy5.5, FITC, and R-PE fluorescence.
      7. PerCP-Cy5.5 is optimized for use with a single argon ion laser emitting 488-nm light. Because of the broad absorption spectrum of the tandem fluorochrome, extra care must be taken when using dual-laser cytometers, which may directly excite both PerCP and Cy5.5™. We recommend the use of cross-beam compensation during data acquisition or software compensation during data analysis.
      8. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
      9. Cy is a trademark of GE Healthcare.
      10. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
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      564133 Rev. 2
      Antibody Details
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      M59-50

      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.  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. 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. 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.

      The M59-50 monoclonal antibody recognizes Stat3 (isoform 1) regardless of phosphorylation status. The specificity of this antibody conjugate for flow cytometric analysis was validated by confirming that RNA-mediated interference (RNAi) of the specific protein reduced the staining of the cells (see figure).  Furthermore, the capacity of the RNAi to down-regulate the expression of the relevant protein was confirmed by western blot analysis.

      564133 Rev. 2
      Format Details
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      PerCP-Cy5.5
      PerCP-Cy5.5 dye is part of the BD blue family of dyes. This tandem fluorochrome is comprised of a fluorescent protein complex (PerCP) with an excitation maximum (Ex Max) of 482 nm and an acceptor dye with an emission maximum (Em Max) at 676 nm. PerCP-Cy5 is designed to be excited by the blue laser (488-nm) and detected using an optical filter centered near 680 nm (e.g., a 695/40 nm bandpass filter). The donor dye can be partially excited by the Violet (405-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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      PerCP-Cy5.5
      Blue 488 nm
      482 nm
      676 nm
      564133 Rev.2
      Citations & References
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      Development References (6)

      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. Darnell JE Jr. STATs and gene regulation. Science. 1997; 277(5332):1630-1635. (Biology). View Reference
      3. Kanai M, Konda Y, Nakajima T, et al . Differentiation-inducing factor-1 (DIF-1) inhibits STAT3 activity involved in gastric cancer cell proliferation via MEK-ERK-dependent pathway. Oncogene. 2003; 22(22):548-554. (Biology). View Reference
      4. Kirito K, Osawa M, Morita H. A functional role of Stat3 in in vivo megakaryopoiesis. Blood. 2002; 99(9):3220-3227. (Biology). View Reference
      5. Redell MS, Ruiz MJ, Gerbing RB, et al. FACS analysis of Stat3/5 signaling reveals sensitivity to G-CSF and IL-6 as a significant prognostic factor in pediatric AML: a Children's Oncology Group report. Blood. 2013; 121(7):1083-1093. (Clone-specific: Flow cytometry). View Reference
      6. Smith PD, Crompton MR. Expression of v-src in mammary epithelial cells induces transcription via STAT3. Biochem J. 1998; 15:331-381. (Biology). View Reference
      View All (6) View Less
      564133 Rev. 2

      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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