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PerCP-Cy™5.5 Rat Anti-Mouse IL-2
PerCP-Cy™5.5 Rat Anti-Mouse IL-2

Flow cytometric analysis for IL-2 in activated mouse splenocytes.  Mouse Intracellular Cytokine-1 positive control cells (MiCK-1) offered by BD Biosciences as MN 554652, are activated mouse splenocytes prepared in the presence of a protein transport inhibitor.  Fixed and permeabilized MiCK-1 cells were stained either with a PerCP-Cy™5.5 Rat IgG2b, κ isotype control (left panel) or with the PerCP-Cy™5.5 Rat Anti-Mouse IL-2 antibody (right panel).  Dot plots were derived from gated events based on light scattering characteristics for lymphocytes.  Flow cytometry was performed on a BD LSR™ II flow cytometry system.

Flow cytometric analysis for IL-2 in activated mouse splenocytes.  Mouse Intracellular Cytokine-1 positive control cells (MiCK-1) offered by BD Biosciences as MN 554652, are activated mouse splenocytes prepared in the presence of a protein transport inhibitor.  Fixed and permeabilized MiCK-1 cells were stained either with a PerCP-Cy™5.5 Rat IgG2b, κ isotype control (left panel) or with the PerCP-Cy™5.5 Rat Anti-Mouse IL-2 antibody (right panel).  Dot plots were derived from gated events based on light scattering characteristics for lymphocytes.  Flow cytometry was performed on a BD LSR™ II flow cytometry system.

Product Details
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BD Pharmingen™
Mouse (QC Testing)
Rat IgG2b
Mouse IL-2 Recombinant Protein
Intracellular staining (flow cytometry) (Routinely Tested)
0.2 mg/ml
AB_1645256
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 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.

Recommended Assay Procedures

Flow cytometry:  The JES6-5H4 antibody is useful for immunofluorescent staining and flow cytometric analysis to identify and enumerate IL-2 producing cells within mixed cell populations.  A useful control investigators may consider using for demonstrating specificity of staining, is to pre-block with one of the following reagents: (1) recombinant mouse IL-2 (Cat. No. 550069) or (2) unlabeled JES6-5H4 antibody (Cat. No. 554425), prior to staining.

Cell Preparation: Investigators not wishing to utilize MiCK-1 cells may alternatively prepare mouse splenocytes (e.g BALB/c) stimulated for 4-6 hours with PMA (5 ng/mL, Sigma-Aldrich Cat. No. P-8139) and ionomycin (500 ng, Sigma-Aldrich Cat. No. I-0634) in the presence of 1 µg/mL Brefeldin A (BD GolgiPlug™ MN 555029).  Investigators are advised to fix and permeabilize the cells prior to staining.

Product Notices

  1. Since applications vary, each investigator should titrate the reagent to obtain optimal results.
  2. An isotype control should be used at the same concentration as the antibody of interest.
  3. 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.
  4. This PerCP-conjugated product is sold under license to the following patent: US Patent No. 4,876,190.
  5. Cy is a trademark of Amersham Biosciences Limited. This conjugated product is sold under license to the following patents: US Patent Nos. 5,486,616; 5,569,587; 5,569,766; 5,627,027.
  6. This product is subject to proprietary rights of Amersham Biosciences Corp. and Carnegie Mellon University and made and sold under license from Amersham Biosciences Corp. This product is licensed for sale only for research. It is not licensed for any other use. If you require a commercial license to use this product and do not have one return this material, unopened to BD Biosciences, 10975 Torreyana Rd, San Diego, CA 92121 and any money paid for the material will be refunded.
  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. 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.
  9. 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.
  10. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
  11. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
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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.

560544 Rev. 1
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
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Citations & References
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Development References (12)

  1. Awatsuji H, Furukawa Y, Nakajima M, Furukawa S, Hayashi K.. Interleukin-2 as a neurotrophic factor for supporting the survival of neurons cultured from various regions of fetal rat brain. J Neurosci Res. 1993; 35(3):305-311. (Biology). View Reference
  2. Fujihashi K, McGhee JR, Beagley KW, et al. Cytokine-specific ELISPOT assay. Single cell analysis of IL-2, IL-4 and IL-6 producing cells. J Immunol Methods. 1993; 160(2):181-189. (Biology). View Reference
  3. Gillis S, Ferm MM, Ou W, Smith KA. T cell growth factor: parameters of production and a quantitative microassay for activity. J Immunol. 1978; 120(6):2027-2032. (Biology). View Reference
  4. Gillis S, Ferm MM, Ou W, Smith KA. T cell growth factor: parameters of production and a quantitative microassay for activity. J Immunol. 1978; 120(6):2027-2032. (Biology). View Reference
  5. Kashima N, Nishi-Takaoka C, Fujita T, et al. Unique structure of murine interleukin-2 as deduced from cloned cDNAs. Nature. 1985; 313(6001):402-404. (Biology). View Reference
  6. Kubo M, Cinader B. Polymorphism of age-related changes in interleukin (IL) production: differential changes of T helper subpopulations, synthesizing IL 2, IL 3 and IL 4. Eur J Immunol. 1990; 20(6):1289-1296. (Biology). View Reference
  7. Mochizuki DY, Watson J, Gillis S. Biochemical separation of interleukin 2. J Immunol Methods. 1980; 39(3):185-201. (Biology). View Reference
  8. Mosmann TR, Cherwinski H, Bond MW, Giedlin MA, Coffman RL. Two types of murine helper T cell clone. I. Definition according to profiles of lymphokine activities and secreted proteins. J Immunol. 1986; 136(7):2348-2357. (Biology). View Reference
  9. 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
  10. 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. (Methodology: ELISA). View Reference
  11. Watson J, Mochizuki D. Interleukin 2: a class of T cell growth factors. Immunol Rev. 1980; 51:257-278. (Biology). View Reference
  12. Yokota T, Arai N, Lee F, Rennick D, Mosmann T, Arai K. Use of a cDNA expression vector for isolation of mouse interleukin 2 cDNA clones: expression of T-cell growth-factor activity after transfection of monkey cells. Proc Natl Acad Sci U S A. 1985; 82(1):68-72. (Biology). View Reference
View All (12) View Less
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For Research Use Only. Not for use in diagnostic or therapeutic procedures.