FITC Rat Anti-Human IL-4
Clone MP4-25D2 (RUO)
- Brand BD Pharmingen™
- Concentration 0.5 mg/ml
- Isotype Rat IgG1
- Reactivity Human (QC Testing)
Intracellular staining (flow cytometry) (Routinely Tested)
- Immunogen Purified Recombinant Human IL-4
- Storage Buffer Aqueous buffered solution containing ≤0.09% sodium azide.
- Regulatory Status RUO
Regulatory Status Legend
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.
This antibody is routinely tested by flow cytometric analysis. Other applications were tested at BD Biosciences Pharmingen during antibody development only or reported in the literature.
FITC, fluorescein isothiocyanate, is a fluorochrome with a molecular weight of 389 Da. FITC is sensitive to pH changes and photobleaching. Due to nearly identical excitation and emission properties but different spillover characteristics, FITC and Alexa Fluor® 488 cannot be used simultaneously. FITC is relatively dim and should be reserved for highly expressed markers whenever possible.
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Preparation and Storage
The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography.
The antibody was conjugated with FITC under optimum conditions, and unreacted FITC was removed.
Store undiluted at 4°C and protected from prolonged exposure to light. Do not freeze.
- Since applications vary, each investigator should titrate the reagent to obtain optimal results.
- Please refer to www.bdbiosciences.com/pharmingen/protocols for technical protocols.
- 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.
Immunofluorescent Staining and Flow Cytometry Analysis: The FITC-conjugated MP4-25D2 antibody can be used for multicolor immunofluorescent staining and flow cytometric analysis to identify and enumerate IL-4-producing cells within mixed cell populations (see image, left panel). For optimal immunofluorescent staining for flow cytometric analysis, this antibody should be titrated (≤ 0.5 µg mAb/million cells). A useful control for demonstrating specificity of staining is the following: pre-block the fixed/permeabilized cells with unlabelled MP4-25D2 antibody (Cat. No. 554482) prior to staining. The intracellular cytokine staining technique and use of blocking controls are described in detail by C. Prussin and D. Metcalfe. A suitable rat IgG1 isotype control for assessing the level of background staining on paraformaldehyde-fixed/saponin-permeabilized human cells is FITC-R3-34 immunoglobulin (Cat. No. 554684); use at comparable concentrations to antibody of interest (e.g. , ≤ 0.5 µg mAb/ 1 million cells). 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 MP4-25D2 (Cat. No. 554483) antibody is useful as a detection antibody for a sandwich ELISA that measures human IL-4 protein levels. Biotinylated MP4-25D2 antibody can be paired with the purified 8D4-8 antibody (Cat. No. 554515) as the capture antibody, with recombinant human IL-4 protein (Cat. No. 554605) as the standard. For assay of IL-4 in serum or plasma, the BD OptEIA™ Human IL-4 ELISA Set (Cat. No. 555194) or BD OptEIA™ Human IL-4 ELISA Kit (Cat. No. 550614) is recommended.
Neutralization: The NA/LE format of the MP4-25D2 antibody (Cat. No. 554481) has been reported to be useful for neutralization of human IL-4 bioactivity. A suitable NA/LE™ rat IgG1 isotype control to match the NA/LE™ MP4-25D2 antibody is the R3-34 antibody, Cat. No. 554683.