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RY586 Rat Anti-Mouse IL-2
RY586 Rat Anti-Mouse IL-2
Two color flow cytometric analysis of IL-2 expression in Stimulated Mouse splenic leucocytes. BALB/c Mouse splenocytes were stimulated for 5 hours with Phorbol 12-Myristate 13-Acetate (PMA; Sigma P-8139; 50 ng/ml) and Ionomycin (Sigma I-0634; 1 μg/ml) in the presence of BD GolgiStop™ Protein Transport Inhibitor (containing Monensin) [Cat. No. 554724]. The cells were fixed with BD Cytofix™ Fixation Buffer (Cat. No. 554655), washed, permeabilized and stained in BD Perm/Wash™ Buffer (Cat. No. 554723) with FITC Rat Anti-Mouse CD4 antibody (Cat. No. 553046/553047) and with either BD Horizon™ RY586 Rat IgG2b, κ Isotype Control (Cat. No. 568160; Left Plot) or BD Horizon™ RY586 Rat Anti-Mouse IL-2 antibody (Cat. No. 568543/568544; Right Plot) at 0.125 µg/test using the BD Biosciences Intracellular Cytokine Staining protocol. The bivariate pseudocolor density plot showing the correlated expression of IL-2 (or Ig Isotype control staining) versus CD4 was derived from gated events with the forward and side-light scatter characteristics of intact stimulated leucocytes. Flow cytometry and data analysis were performed using a BD LSRFortessa™ X-20 Cell Analyzer System and FlowJo™ software. Data shown on this Technical Data Sheet are not lot specific.
Two color flow cytometric analysis of IL-2 expression in Stimulated Mouse splenic leucocytes. BALB/c Mouse splenocytes were stimulated for 5 hours with Phorbol 12-Myristate 13-Acetate (PMA; Sigma P-8139; 50 ng/ml) and Ionomycin (Sigma I-0634; 1 μg/ml) in the presence of BD GolgiStop™ Protein Transport Inhibitor (containing Monensin) [Cat. No. 554724]. The cells were fixed with BD Cytofix™ Fixation Buffer (Cat. No. 554655), washed, permeabilized and stained in BD Perm/Wash™ Buffer (Cat. No. 554723) with FITC Rat Anti-Mouse CD4 antibody (Cat. No. 553046/553047) and with either BD Horizon™ RY586 Rat IgG2b, κ Isotype Control (Cat. No. 568160; Left Plot) or BD Horizon™ RY586 Rat Anti-Mouse IL-2 antibody (Cat. No. 568543/568544; Right Plot) at 0.125 µg/test using the BD Biosciences Intracellular Cytokine Staining protocol. The bivariate pseudocolor density plot showing the correlated expression of IL-2 (or Ig Isotype control staining) versus CD4 was derived from gated events with the forward and side-light scatter characteristics of intact stimulated leucocytes. Flow cytometry and data analysis were performed using a BD LSRFortessa™ X-20 Cell Analyzer System and FlowJo™ software. Data shown on this Technical Data Sheet are not lot specific.
Product Details
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BD Horizon™
Il2; Interleukin-2; T-cell growth factor; TCGF
Mouse (QC Testing)
Rat IgG2b
Mouse IL-2 Recombinant Protein
Intracellular staining (flow cytometry) (Routinely Tested)
0.2 mg/ml
16183
Aqueous buffered solution containing ≤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 the dye under optimum conditions and unreacted dye was removed.

Recommended Assay Procedures

BD® CompBeads can be used as surrogates to assess fluorescence spillover (compensation). When fluorochrome conjugated antibodies are bound to BD® CompBeads, they have spectral properties very similar to cells. However, for some fluorochromes there can be small differences in spectral emissions compared to cells, resulting in spillover values that differ when compared to biological controls. It is strongly recommended that when using a reagent for the first time, users compare the spillover on cells and BD® CompBeads to ensure that BD® CompBeads are appropriate for your specific cellular application.

Product Notices

  1. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
  2. 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.
  3. Since applications vary, each investigator should titrate the reagent to obtain optimal results.
  4. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
  5. An isotype control should be used at the same concentration as the antibody of interest.
  6. CF™ is a trademark of Biotium, Inc.
  7. Please refer to http://regdocs.bd.com to access safety data sheets (SDS).
  8. For U.S. patents that may apply, see bd.com/patents.
568544 Rev. 1
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.

568544 Rev. 1
Format Details
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RY586
The BD Horizon RealYellow™ 586 (RY586) Dye is part of the BD family of yellow-green dyes. It is a small organic fluorochrome with an excitation maximum (Ex Max) at 565-nm and an emission maximum (Em Max) at 586-nm. Driven by BD innovation, RY586 can be used on both spectral and conventional cytometers and is designed to be excited by the Yellow-Green laser (561-nm) with minimal excitation by the 488-nm Blue laser. For conventional instruments equipped with a Yellow-Green laser (561-nm), RY586 can be used as an alternative to PE and we recommend using an optical filter centered near 586-nm (eg, a 586/15-nm bandpass filter). For spectral instruments equipped with a Yellow-Green laser (561-nm), it can be used in conjunction with PE. Compared to PE, RY586 is similar in brightness, minimal spillover into Blue detectors, and increased spillover into the 610/20-nm (PE-CF594) detector. Please ensure that your instrument configuration (lasers and optical filters) is appropriate for this dye.
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RY586
Yellow-Green 561 nm
564 nm
586 nm
568544 Rev.1
Citations & References
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Development References (5)

  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. Karulin AY, Hesse MD, Tary-Lehmann M, Lehmann PV. Single-cytokine-producing CD4 memory cells predominate in type 1 and type 2 immunity.. J Immunol. 2000; 164(4):1862-72. (Clone-specific: Flow cytometry). View Reference
  4. 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
  5. 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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568544 Rev. 1

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