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      Alexa Fluor® 647 Mouse Anti-Human IFN-γ
      Alexa Fluor® 647 Mouse Anti-Human IFN-γ
      Two-color flow cytometric analysis of IFN- γ expression in stimulated human peripheral blood mononuclear cells (PBMC). Human PBMC were stimulated for 6 hours with Phorbol 12-Myristate 13-Acetate (Sigma  P-8139; 50 ng/ml final concentration) and Ionomycin (Sigma I-0634; 1 μg/ml final concentration) in the presence of BD GolgiStop™ (Cat. No. 554724). The PBMC were fixed and permeabilized with BD Cytofix/Cytoperm™ Fixation and Permeabilization Solution (Cat. No. 554722) and washed with BD Perm/Wash™ Buffer (Cat. No. 554723). The cells were then stained with PE Mouse Anti-Human CD3 antibody (Cat.No. 555333/561808/561809) and either Alexa Fluor® 647 Mouse IgG1 κ Isotype Control (Cat. No. 557732; Left Panel) or Alexa Fluor® 647 Mouse Anti-Human IFN-γ antibody (Cat. No. 563495; Right Panel). Two-color flow cytometric dot plots show the correlated expression patterns of IFN-γ (or Ig Isotype control staining) versus CD3 for gated events with the forward and side light-scatter characteristics of intact PBMC. Flow cytometric analysis was performed using a BD™ LSR II Flow Cytometer System.
      Alexa Fluor® 647 Mouse Anti-Human IFN-γ
      Two-color flow cytometric analysis of IFN- γ expression in stimulated human peripheral blood mononuclear cells (PBMC). Human PBMC were stimulated for 6 hours with Phorbol 12-Myristate 13-Acetate (Sigma  P-8139; 50 ng/ml final concentration) and Ionomycin (Sigma I-0634; 1 μg/ml final concentration) in the presence of BD GolgiStop™ (Cat. No. 554724). The PBMC were fixed and permeabilized with BD Cytofix/Cytoperm™ Fixation and Permeabilization Solution (Cat. No. 554722) and washed with BD Perm/Wash™ Buffer (Cat. No. 554723). The cells were then stained with PE Mouse Anti-Human CD3 antibody (Cat.No. 555333/561808/561809) and either Alexa Fluor® 647 Mouse IgG1 κ Isotype Control (Cat. No. 557732; Left Panel) or Alexa Fluor® 647 Mouse Anti-Human IFN-γ antibody (Cat. No. 563495; Right Panel). Two-color flow cytometric dot plots show the correlated expression patterns of IFN-γ (or Ig Isotype control staining) versus CD3 for gated events with the forward and side light-scatter characteristics of intact PBMC. Flow cytometric analysis was performed using a BD™ LSR II Flow Cytometer System.
      Product Details
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      BD Pharmingen™
      IFNG; Interferon-gamma; IFG; IFI; Type II interferon
      Human (QC Testing), Rhesus, Cynomolgus, Baboon (Tested in Development)
      Mouse BALB/c IgG1, κ
      Human IFN-γ from supernatants of S. aureus-stimulated PBMC
      Intracellular staining (flow cytometry) (Routinely Tested)
      5 µl
      3458
      AB_2738241
      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 to Alexa Fluor® 647 under optimum conditions, and unreacted Alexa Fluor® 647 was removed.
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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. An isotype control should be used at the same concentration as the antibody of interest.
      4. The Alexa Fluor®, Pacific Blue™, and Cascade Blue® dye antibody conjugates in this product are sold under license from Molecular Probes, Inc. for research use only, excluding use in combination with microarrays, or as analyte specific reagents. The Alexa Fluor® dyes (except for Alexa Fluor® 430), Pacific Blue™ dye, and Cascade Blue® dye are covered by pending and issued patents.
      5. Alexa Fluor® is a registered trademark of Molecular Probes, Inc., Eugene, OR.
      6. Alexa Fluor® 647 fluorochrome emission is collected at the same instrument settings as for allophycocyanin (APC).
      7. 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.
      8. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
      9. Species cross-reactivity detected in product development may not have been confirmed on every format and/or application.
      10. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
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      563495 Rev. 2
      Antibody Details
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      4S.B3

      The 4S.B3 monoclonal antibody specifically binds to interferon-γ (IFN-γ). The immunogen used to generate this hybridoma was partially purified human IFN-γ obtained from supernatants of human PBMC stimulated with Staphylococcus aureus. Interferon-γ (IFN-γ) is a potent multifunctional cytokine that is produced by several activated cell types including NK, NKT, CD4+TCRαβ+, CD8+TCRαβ+, and TCRγδ+ T cells. IFN-γ exerts its biological effects through specific binding to the high-affinity IFN-γ Receptor Complex comprised of IFN-γRα (CD119) and IFN-γRβ subunits. In addition to its antiviral effects, IFN-γ upregulates a number of lymphoid cell functions including the antimicrobial and antitumor responses of macrophages, NK cells, and neutrophils. In addition, IFN-γ can exert strong regulatory influences on the proliferation, differentiation, and effector responses of B cell and T cell subsets. These influences can involve IFN-γ's capacity to boost MHC class I and II expression by antigen-presenting cells as well as to direct effects on B cells and T cells themselves. Human IFN-γ is a 14-18 kDa glycoprotein containing 143 amino acid residues.

      Clone 4S.B3 also cross-reacts with a cytoplasmic component of peripheral blood CD3+ lymphocytes of baboon, and both rhesus and cynomolgus macaque monkeys following five-hour treatment with phorbol myristic acetate (PMA) and Ca++ ionophore (A23187) in the presence of monensin. The staining pattern of 4S.B3 in CD3+ cells is similar to that observed with peripheral blood T lymphocytes from normal human donors. This reagent is useful for intracellular immunofluorescent staining for flow cytometric analysis to identify and enumerate IFN-γ + cells within a mixed cell population.

      563495 Rev. 2
      Format Details
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      Alexa Fluor™ 647
      Alexa Fluor™ 647 Dye is part of the BD red family of dyes. This is a small organic fluorochrome with an excitation maximum (Ex Max) at 653-nm and an emission maximum (Em Max) at 669-nm. Alexa Fluor 647 is designed to be excited by the Red laser (627-640 nm) and detected using an optical filter centered near 520-nm (e.g., a 660/20 nm bandpass filter). Please ensure that your instrument’s configurations (lasers and optical filters) are appropriate for this dye.
      Alexa Fluor™ 647
      Red 627-640 nm
      653 nm
      669 nm
      563495 Rev.2
      Citations & References
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      View product citations for antibody "563495" on CiteAb

      Development References (5)

      1. Meager A, Parti S, Barwick S, Spragg J, O'Hagan K. Detection of hybridomas secreting monoclonal antibodies to human gamma interferon using a rapid screening technique and specificity of certain monoclonal antibodies to gamma interferon. J Interferon Res. 1984; 4(4):619-625. (Immunogen: Immunoprecipitation, Radioimmunoassay). View Reference
      2. Meager A. Characterization of interferons and immunoassays. In: Clemens MJ, Morris AG, Gearing AJH, ed. Lymphokines and Interferons. A Practical Approach. Oxford: IRL Press Ltd; 1987:105-127.
      3. Prussin C, Metcalfe DD. Detection of intracytoplasmic cytokine using flow cytometry and directly conjugated anti-cytokine antibodies. J Immunol Methods. 1995; 188(1):117-128. (Methodology: Flow cytometry). View Reference
      4. Rotteveel FT, Kokkelink I, van Lier RA, et al. Clonal analysis of functionally distinct human CD4+ T cell subsets. J Exp Med. 1988; 168(5):1659-1673. (Clone-specific: ELISA). View Reference
      5. Vogel S, Friedman R, Hogan M. Measurement of antiviral activity induced by interferons a, b, and g.. In: Coligan JE, Kruisbeek AM, Margulies DH, Shevach EM, Strober W, ed. Current Protocols in Immunology. New York: John Wiley & Sons; 2007:6.9.1-6.9.15.
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      563495 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.