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      Purified Rat Anti-Mouse IL-2
      Purified Rat Anti-Mouse IL-2
      Expression of IL-2 by stimulated CD4+ and CD4- BALB/c spleen cells. Splenocytes from 6 month old BALB/c mice were stimulated for 5 hours with hamster anti-mouse CD3 (2 µg/ml final concentration; clone 145-2C11, Cat. No. 553057) and hamster anti-mouse CD28 (2 µg/ml final concentration; clone 37.51, Cat. No. 553294) antibodies in the presence of BD GolgiStop™ (3 µM final concentration; Cat. No. 554724). The cells were harvested, stained with 0.06 µg of PE-conjugated rat anti-mouse CD4 (PE-RM4-5, Cat. No. 553049), fixed, permeabilized, and subsequently stained with 0.06 µg of FITC-conjugated rat anti-mouse IL-2 antibody (FITC-JES6-5H4, Cat. No. 554427, left panel). To demonstrate specificity of staining, the binding by the FITC-JES6-5H4 antibody was blocked by preincubation of the conjugated antibody with recombinant mouse IL-2 (0.12 µg, Cat. No. 550069; middle panel), and by preincubation of the fixed/permeabilized cells with unlabeled JES6-5H4 antibody (2.5 µg; Cat. No. 554425; right panel) prior to staining. The quadrant markers for the bivariate dot plots were set based on the autofluorescence controls and verified using the recombinant cytokine blocking and unlabelled antibody blocking specificity controls.
      Purified Rat Anti-Mouse IL-2
      Expression of IL-2 by stimulated CD4+ and CD4- BALB/c spleen cells. Splenocytes from 6 month old BALB/c mice were stimulated for 5 hours with hamster anti-mouse CD3 (2 µg/ml final concentration; clone 145-2C11, Cat. No. 553057) and hamster anti-mouse CD28 (2 µg/ml final concentration; clone 37.51, Cat. No. 553294) antibodies in the presence of BD GolgiStop™ (3 µM final concentration; Cat. No. 554724). The cells were harvested, stained with 0.06 µg of PE-conjugated rat anti-mouse CD4 (PE-RM4-5, Cat. No. 553049), fixed, permeabilized, and subsequently stained with 0.06 µg of FITC-conjugated rat anti-mouse IL-2 antibody (FITC-JES6-5H4, Cat. No. 554427, left panel). To demonstrate specificity of staining, the binding by the FITC-JES6-5H4 antibody was blocked by preincubation of the conjugated antibody with recombinant mouse IL-2 (0.12 µg, Cat. No. 550069; middle panel), and by preincubation of the fixed/permeabilized cells with unlabeled JES6-5H4 antibody (2.5 µg; Cat. No. 554425; right panel) prior to staining. The quadrant markers for the bivariate dot plots were set based on the autofluorescence controls and verified using the recombinant cytokine blocking and unlabelled antibody blocking specificity controls.
      Product Details
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      BD Pharmingen™
      Mouse (QC Testing)
      Rat IgG2b
      Mouse IL-2 Recombinant Protein
      Intracellular block/flow cytometry (Routinely Tested), ELISA (Tested During Development), Immunoprecipitation (Reported)
      0.5 mg/ml
      16183
      AB_398554
      Aqueous buffered solution containing ≤0.09% sodium azide.
      RUO


      Preparation And Storage

      The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography. Store undiluted at 4°C.
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      Recommended Assay Procedures

      Recommended Assay Procedure:

      Blocking Control for Intracellular Staining: The purified JES6-5H4 antibody can be used as a blocking control to demonstrate specificity of IL-2 staining by directly conjugated-JES6-5H4 antibodies. To perform this control, the fixed/permeabilized cells (~ 1 million) can be incubated with 1-10 µg of unlabeled JES6-5H4 antibody (Cat. No. 554425) for 20 minutes at 4°C, prior to staining with conjugated-JES6-5H4 (e.g., 0.1 -0.5 µg mAb/1 million cells). The intracellular cytokine staining technique and use of blocking controls are described in detail by C. Prussin and D. Metcalfe. For specific methodology, please visit our web site, www.bdbiosciences.com, and go to the protocols section or the chapter on intracellular staining in the Immune Function Handbook.

      ELISA Detection: The biotinylated JES6-5H4 antibody (Cat. No. 554426) is useful as a detection antibody for a sandwich ELISA for measuring mouse IL-2 protein levels.

      IP: The purified JES6-5H4 antibody has been reported useful for immunoprecipitation. Please note that this application is not routinely tested at BD Biosciences Pharmingen.

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      Product Notices

      1. Since applications vary, each investigator should titrate the reagent to obtain optimal results.
      2. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
      3. 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.
      4. Use of these products to measure activation antigens expressed on mononuclear cell subsets for the purpose of monitoring immunoregulatory status can fall under one or more claims of the following patents: US Patent Nos. 5,445,939, 5,656,446, 5,843,689; European Patent No. 319,543; Canadian Patent No. 1,296,622; Australian Patent No. 615,880; and Japanese Patent No. 2,769,156.
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      554425 Rev. 3
      Antibody Details
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      JES6-5H4

      The JES6-5H4 monoclonal antibody specifically binds to mouse interleukin-2 (IL-2), a multifunctional cytokine that plays pivotal roles in immunity and tolerance. It is produced by activated T cells and affects the activation, growth, proliferation and/or differentiation of various cell types including T and B lymphocytes and their precursors, LAK cells, NK cells, and monocytes/macrophages. IL-2 mediates its biological activities by binding to IL-2 receptor complexes. The intermediate affinity IL-2R is comprised of IL-2Rβ (CD122) and common gamma chain (γc; CD132) subunits, whereas the high-affinity IL-2R is comprised of IL-2Rα (CD25), IL-2Rβ, and γc subunits. The JES6-5H4 monoclonal antibody binds to IL-2 and neutralizes its biological activity.

      554425 Rev. 3
      Format Details
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      Purified
      Tissue culture supernatant is purified by either protein A/G or affinity purification methods. Both methods yield antibody in solution that is free of most other soluble proteins, lipids, etc. This format provides pure antibody that is suitable for a number of downstream applications including: secondary labeling for flow cytometry or microscopy, ELISA, Western blot, etc.
      Purified
      554425 Rev.3
      Citations & References
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      View product citations for antibody "554425" on CiteAb

      Development References (4)

      1. Abrams J. Immunoenzymetric assay of mouse and human cytokines using NIP-labeled anti-cytokine antibodies. Curr Protoc Immunol. 2001; 1:6.20-6.21. (Clone-specific: ELISA). View Reference
      2. Abrams JS, Roncarolo MG, Yssel H, Andersson U, Gleich GJ, Silver JE. Strategies of anti-cytokine monoclonal antibody development: immunoassay of IL-10 and IL-5 in clinical samples. Immunol Rev. 1992; 127:5-24. (Clone-specific: ELISA, Immunoprecipitation). View Reference
      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. Sander B, Hoiden I, Andersson U, Moller E, Abrams JS. Similar frequencies and kinetics of cytokine producing cells in murine peripheral blood and spleen. Cytokine detection by immunoassay and intracellular immunostaining. J Immunol Methods. 1993; 166(2):201-214. (Clone-specific: ELISA). View Reference
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      554425 Rev. 3

       

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      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.