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Expression of IL-4 by stimulated human peripheral blood mononuclear cells. Human peripheral blood mononuclear cells were stimulated with immobilized Purified NA/LE Mouse Anti-Human CD3 (Cat. No. 555329; 10 µg/ml for plate coating) and soluble Purified NA/LE Mouse Anti-Human CD28 (Cat. No. 555725; 2 µg/ml) antibodies, recombinant human IL-2 (10 ng/ml; Cat. No. 554603) and human IL-4 (25 ng/ml; Cat. No. 554605) for 2 days. The cells were subsequently cultured in medium containing recombinant human IL-2 and human IL-4 for 3 days. Finally, the cells were harvested and stimulated for 5 hr with Phorbol 12-Myristate 13-Acetate (Sigma, Cat. No. P-8139; 50 ng/ml) and Ionomycin (Sigma, Cat. No. I-0634; 1 μg/ml) in the presence of BD GolgiStop™ Protein Transport Inhibitor (containing Monensin) (Cat. No. 554724). The cells were harvested, washed with BD Pharmingen™ Stain Buffer (FBS) (Cat. No. 554656), and fixed and permeabilized with BD Cytofix/Cytoperm™ Fixation and Permeabilization Solution (554722). The cells were then washed and stained in BD Perm/Wash™ Buffer (Cat. No. 554723) with FITC Mouse Anti-Human CD4 (Cat. No. 555346/561005/561842) and either BD Horizon™ BV421 Rat IgG1 κ Isotype Control (Cat. No. 562868; Left Panel) or BD Horizon™ BV421 Mouse Anti-Human IL-4 antibody (Cat. No. 564110; Right Panel) using BD Biosciences Intracellular Cytokine Staining protocol. Two-color flow cytometric dot plots show the correlated expression patterns of IL-4 (or Ig Isotype control staining) versus CD4 for gated events with the forward and side light-scatter characteristics of intact stimulated lymphocytes. Flow cytometric analysis was performed using a BD™ LSR II Flow Cytometer System.


BD Horizon™ BV421 Rat Anti-Human IL-4

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Any use of products other than the permitted use without the express written authorization of Becton, Dickinson and Company is strictly prohibited.
Preparation And Storage
Product Notices
- 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).
- An isotype control should be used at the same concentration as the antibody of interest.
- 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.
- Source of all serum proteins is from USDA inspected abattoirs located in the United States.
- Pacific Blue™ is a trademark of Molecular Probes, Inc., Eugene, OR.
- Brilliant Violet™ 421 is a trademark of Sirigen.
- For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
- Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
Companion Products






The MP4-25D2 monoclonal antibody specifically binds to Interleukin-4 (IL-4). IL-4 is also known as Lymphocyte stimulatory factor 1, B cell stimulatory factor 1 (BSF-1), or B cell growth factor 1 (BCGF-1). IL-4 is produced by activated T cells, basophils, and mast cells. IL-4 is a multifunctional cytokine and growth factor that affects a variety of different target cell types. IL-4 can costimulate T cell proliferation and survival, as well as help drive Th2-like cell differentiation and effector functions. It costimulates B cell proliferation and survival, and can help B cells differentiate into IgG4- or IgE-producing cells. IL-4 can also diminish the proinflammatory functions of monocytes and macrophages. IL-4 exerts its biological effects by binding with high affinity to cell surface CD124 (IL-4Rα chain). CD124 forms signaling IL-4 receptor complexes with either CD132/common γ chain or CD213a1/IL-13Rα1 to form either Type I or Type II IL-4 Receptor complexes, respectively. The immunogen used to generate the MP4-25D2 hybridoma was purified recombinant human IL-4. This is a neutralizing antibody. The MP4-25D2 antibody has been reported to crossreact with IL-4 from rhesus monkeys. The use of the MP4-25D2 antibody for epitope mapping of human IL-4 has been described.
The antibody was conjugated to BD Horizon™ BV421 which is part of the BD Horizon™ Brilliant Violet™ family of dyes. With an Ex Max of 407-nm and Em Max at 421-nm, BD Horizon™ BV421 can be excited by the violet laser and detected in the standard Pacific Blue™ filter set (eg, 450/50-nm filter). BD Horizon™ BV421 conjugates are very bright, often exhibiting a 10 fold improvement in brightness compared to Pacific Blue™ conjugates.

Development References (5)
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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, Neutralization). View Reference
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Chretien I, Van Kimmenade A, Pearce MK, Banchereau J, Abrams JS. Development of polyclonal and monoclonal antibodies for immunoassay and neutralization of human interleukin-4. J Immunol Methods. 1989; 117(1):67-71. (Clone-specific: ELISA, Neutralization). View Reference
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Jung T, Schauer U, Rieger C, et al. Interleukin-4 and interleukin-5 are rarely co-expressed by human T cells. Eur J Immunol. 1995; 25(8):2413-2416. (Clone-specific: Flow cytometry). View Reference
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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: IC/FCM Block). View Reference
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Ramanathan L, Ingram R, Sullivan L, et al. Immunochemical mapping of domains in human interleukin 4 recognized by neutralizing monoclonal antibodies. Biochemistry. 1993; 32(14):3549-3556. (Clone-specific: Functional assay, Neutralization). View Reference
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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.