PE Rat Anti-Human IL-13

BD Pharmingen™ PE Rat Anti-Human IL-13

Name: PE Rat Anti-Human IL-13
Brand: BD Pharmingen™
SKU: 562039

Clone JES10-5A2

(RUO)

Expression of IL-13 by stimulated human CD4+ cells. Isolated human CD4+ cells were stimulated with immobilized anti-human CD3 antibody (UCHT1, 10 µg/ml for plate coating, Cat. No. 555329), soluble anti-human CD28 antibody (20 ng/ml, Cat. No. 555725), recombinant human IL-2 (10 ng/ml, Cat. No. 554603) and recombinant human IL-4 (10 ng/ml, Cat No. 554605) for 2 days. The cells were washed and subsequently cultured in medium containing IL-2 and IL-4 for 3 days. Finally, the cells were harvested and re-stimulated for 6 hr with PMA (Sigma, Cat. #P-8139; 50 ng/ml), calcium ionophore A23187 (Sigma, Cat. #C-9275; 250 ng/ml) in the presence of GolgiStop™ (2 µM final concentration, Cat. No. 554724). The cells were stained with PE-Cy™5 anti-CD4 (Cat No. 555348) fixed, permeabilized, and subsequently stained with 0.25 µg of PE-rat anti-human IL-13 antibody (Cat No. 562039/554571) by using Pharmingen's staining protocol (see left panel). To demonstrate specificity of staining, the binding of PE rat anti-human IL-13 antibody was blocked by the preincubation of the conjugated antibody with excess recombinant human IL-13 (see middle panel), and by preincubation of the fixed/permeabilized cells with Purified Rat Anti-Human IL-13 antibody (5 µg, Cat No. 554570; see right panel) prior to staining with the PE rat anti-human IL-13 antibody. The quadrant markers for the bivariate dot plots were set based on the autofluorescence control and verified with the recombinant cytokine blocking and unlabeled antibody blocking specificity controls.

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Product Details

Brand: BD Pharmingen™

Alternative Name: Interleukin-13; NC30; ALRH; BHR1; P600

Reactivity: Human (QC Testing)

Isotype: Rat IgG1

Immunogen: Human Recombinant IL-13

Application: Intracellular staining (flow cytometry) (Routinely Tested)

Concentration: 0.2 mg/ml

Entrez Gene ID: 3596

RRID: AB_395485

Storage Buffer: Aqueous buffered solution containing ≤0.09% sodium azide.

Regulatory Status: 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 unconjugated antibody and free dye were removed.

Recommended Assay Procedures

Immunofluorescent Staining and Flow Cytometric Analysis: The PE Rat Anti-Human IL-13 (Cat. No. 554571) can be used for multicolor immunofluorescent staining and flow cytometric analysis to identify and enumerate IL-13 producing cells within mixed cell populations (see Figure). For optimal immunofluorescent staining and flow cytometric analysis, this anti-cytokine antibody should be titrated (≤ 0.5 µg mAb/million cells). For specific methodology, please visit our website, https://www.bdbiosciences.com/en-us/resources/protocols.

A useful control for demonstrating specificity of staining is either of the following: 1) pre-block the PE Rat Anti-Human IL-13 antibody with ligand (e.g., human IL-13) prior to staining, or 2) pre-block paraformaldehyde-fixed/saponin-permeabilized cells with unlabeled Rat Anti-Human IL-13 antibody (e.g., Cat. No. 554570) prior to staining. The intracellular staining technique and use of blocking controls are described in detail by C. Prussin and D. Metcalfe. A suitable rat IgG1 isotype control immunoglobulin for assessing the level of background staining on paraformaldehyde-fixed/saponin-permeabilized mouse or human cells is the PE Rat IgG1, κ Isotype Control (Cat. No. 554685); use at comparable concentrations to antibody of interest (e.g., ≤ 0.5 µg mAb/1 million cells).

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 cell and BD® CompBeads to ensure that BD® CompBeads are appropriate for your specific cellular application.

Product Notices

Since applications vary, each investigator should titrate the reagent to obtain optimal results.
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.
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.
Please refer to http://regdocs.bd.com to access safety data sheets (SDS).

562039 Rev. 1

Antibody Details

JES10-5A2

The JES10-5A2 monoclonal antibody specifically binds to human interleukin-13, IL-13. IL-13 is produced by activated T cells, mast cells and NK cells. IL-13 regulates IgE production by B cells and can suppress the cytotoxic activity of macrophages and their production of inflammatory mediators. The immunogen used to produce the JES10-5A2 hybridoma was COS-expressed recombinant human IL-13. This is a neutralizing antibody.

562039 Rev. 1

Format Details

R-Phycoerythrin (PE), is part of the BD family of Phycobiliprotein dyes. This fluorochrome is a multimeric fluorescent phycobiliprotein with excitation maximum (Ex Max) of 496 nm and 566 nm and an emission maximum (Em Max) at 576 nm. PE is designed to be excited by the Blue (488 nm), Green (532 nm) and Yellow-Green (561 nm) lasers and detected using an optical filter centered near 575 nm (e.g., a 575/26-nm bandpass filter). As PE is excited by multiple lasers, this can result in cross-laser excitation and fluorescence spillover on instruments with various combinations of Blue, Green, and Yellow-Green lasers. Please ensure that your instrument’s configurations (lasers and optical filters) are appropriate for this dye.

Format: PE

Excitation Source: Yellow-Green 488 nm, 532 nm, 561 nm

Excitation Max: 496 nm, 566 nm

Emission Max: 576 nm

562039 Rev.1

Citations & References

Development References (3)

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
McKenzie A, Zurawski G. Measurement of IL-13. In: Coligan, Kruisbeek, Shevak, Strober, ed. Current Protocols in Immunology. New York: John Wiley & Sons; 1994:18-19.
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

562039 Rev. 1

Please refer to Support Documents for Quality Certificates

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.