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BD Horizon™ BV711 Rat Anti-Mouse IL-10
Clone JES5-16E3 (RUO)
Two-color flow cytometric analysis of IL-10 expression in stimulated mouse T cells. Splenic CD4+ T cells were cultured (2 days) with plate-bound Purified NA/LE Hamster Anti-Mouse CD3e (Cat. No. 553057; 25 μg/ml for coating) and soluble Purified NA/LE Hamster Anti-Mouse CD28 (Cat. No. 553294; 2 μg/ml) antibodies and with Recombinant IL-2 (Cat. No. 550069; 10 ng/ml) and IL-4 (Cat. No. 550067; 25 ng/ml). The stimulated cells were subsequently cultured (3 days) with Recombinant IL-2 and IL-4. The cells were then stimulated with PMA (Sigma Cat. No. P-8139) and Ionomycin (Sigma Cat. No. I-0634) in the presence of BD GolgiStop™ Protein Transport Inhibitor (containing Monensin) (Cat. No. 554724). After 6 hours, the cells were harvested, washed, fixed and permeabilized with BD Cytofix/Cytoperm™ Fixation and Permeabilization Solution (Cat. No. 554722).
The cells were then washed and stained in BD Perm/Wash™ Buffer (Cat. No. 554723) with BD Horizon™ BV711 Rat IgG2b, κ Isotype Control (Cat. No. 563045; Left Panel) or BD Horizon BV711 Rat Anti-Mouse IL-10 antibody (Cat. No. 564081; Right Panel). Flow cytometric contour plots showing correlated expression of IL-10 (or Ig Isotype control staining) versus Autofluorescence were derived from gated events with the forward and side light-scatter characteristics of intact stimulated lymphocytes. Flow cytometric analysis was performed using a BD LSRFortessa™ Cell Analyzer System.
Two-color flow cytometric analysis of IL-10 expression in stimulated mouse T cells. Splenic CD4+ T cells were cultured (2 days) with plate-bound Purified NA/LE Hamster Anti-Mouse CD3e (Cat. No. 553057; 25 μg/ml for coating) and soluble Purified NA/LE Hamster Anti-Mouse CD28 (Cat. No. 553294; 2 μg/ml) antibodies and with Recombinant IL-2 (Cat. No. 550069; 10 ng/ml) and IL-4 (Cat. No. 550067; 25 ng/ml). The stimulated cells were subsequently cultured (3 days) with Recombinant IL-2 and IL-4. The cells were then stimulated with PMA (Sigma Cat. No. P-8139) and Ionomycin (Sigma Cat. No. I-0634) in the presence of BD GolgiStop™ Protein Transport Inhibitor (containing Monensin) (Cat. No. 554724). After 6 hours, the cells were harvested, washed, fixed and permeabilized with BD Cytofix/Cytoperm™ Fixation and Permeabilization Solution (Cat. No. 554722).
The cells were then washed and stained in BD Perm/Wash™ Buffer (Cat. No. 554723) with BD Horizon™ BV711 Rat IgG2b, κ Isotype Control (Cat. No. 563045; Left Panel) or BD Horizon BV711 Rat Anti-Mouse IL-10 antibody (Cat. No. 564081; Right Panel). Flow cytometric contour plots showing correlated expression of IL-10 (or Ig Isotype control staining) versus Autofluorescence were derived from gated events with the forward and side light-scatter characteristics of intact stimulated lymphocytes. Flow cytometric analysis was performed using a BD LSRFortessa™ Cell Analyzer System.
Two-color flow cytometric analysis of IL-10 expression in stimulated mouse T cells. Splenic CD4+ T cells were cultured (2 days) with plate-bound Purified NA/LE Hamster Anti-Mouse CD3e (Cat. No. 553057; 25 μg/ml for coating) and soluble Purified NA/LE Hamster Anti-Mouse CD28 (Cat. No. 553294; 2 μg/ml) antibodies and with Recombinant IL-2 (Cat. No. 550069; 10 ng/ml) and IL-4 (Cat. No. 550067; 25 ng/ml). The stimulated cells were subsequently cultured (3 days) with Recombinant IL-2 and IL-4. The cells were then stimulated with PMA (Sigma Cat. No. P-8139) and Ionomycin (Sigma Cat. No. I-0634) in the presence of BD GolgiStop™ Protein Transport Inhibitor (containing Monensin) (Cat. No. 554724). After 6 hours, the cells were harvested, washed, fixed and permeabilized with BD Cytofix/Cytoperm™ Fixation and Permeabilization Solution (Cat. No. 554722).
The cells were then washed and stained in BD Perm/Wash™ Buffer (Cat. No. 554723) with BD Horizon™ BV711 Rat IgG2b, κ Isotype Control (Cat. No. 563045; Left Panel) or BD Horizon BV711 Rat Anti-Mouse IL-10 antibody (Cat. No. 564081; Right Panel). Flow cytometric contour plots showing correlated expression of IL-10 (or Ig Isotype control staining) versus Autofluorescence were derived from gated events with the forward and side light-scatter characteristics of intact stimulated lymphocytes. Flow cytometric analysis was performed using a BD LSRFortessa™ Cell Analyzer System.
Two-color flow cytometric analysis of IL-10 expression in stimulated mouse T cells. Splenic CD4+ T cells were cultured (2 days) with plate-bound Purified NA/LE Hamster Anti-Mouse CD3e (Cat. No. 553057; 25 μg/ml for coating) and soluble Purified NA/LE Hamster Anti-Mouse CD28 (Cat. No. 553294; 2 μg/ml) antibodies and with Recombinant IL-2 (Cat. No. 550069; 10 ng/ml) and IL-4 (Cat. No. 550067; 25 ng/ml). The stimulated cells were subsequently cultured (3 days) with Recombinant IL-2 and IL-4. The cells were then stimulated with PMA (Sigma Cat. No. P-8139) and Ionomycin (Sigma Cat. No. I-0634) in the presence of BD GolgiStop™ Protein Transport Inhibitor (containing Monensin) (Cat. No. 554724). After 6 hours, the cells were harvested, washed, fixed and permeabilized with BD Cytofix/Cytoperm™ Fixation and Permeabilization Solution (Cat. No. 554722).
The cells were then washed and stained in BD Perm/Wash™ Buffer (Cat. No. 554723) with BD Horizon™ BV711 Rat IgG2b, κ Isotype Control (Cat. No. 563045; Left Panel) or BD Horizon BV711 Rat Anti-Mouse IL-10 antibody (Cat. No. 564081; Right Panel). Flow cytometric contour plots showing correlated expression of IL-10 (or Ig Isotype control staining) versus Autofluorescence were derived from gated events with the forward and side light-scatter characteristics of intact stimulated lymphocytes. Flow cytometric analysis was performed using a BD LSRFortessa™ Cell Analyzer System.
Product Details
BD Horizon™
Interleukin-10; Il10; CSIF; Cytokine synthesis inhibitory factor
Mouse (QC Testing)
Rat IgG2b
Recombinant mouse IL-10
Intracellular staining (flow cytometry) (Routinely Tested)
0.2 mg/ml
AB_2738581
Aqueous buffered solution containing BSA and ≤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 BD Horizon™ BV711 under optimum conditions, and unconjugated antibody and free BD Horizon™ BV711 were removed.
Recommended Assay Procedures
For optimal and reproducible results, BD Horizon Brilliant™ Stain Buffer should be used anytime BD Horizon Brilliant™ dyes are used in a multicolor flow cytometry panel. Fluorescent dye interactions may cause staining artifacts which may affect data interpretation. The BD Horizon Brilliant Stain Buffer was designed to minimize these interactions. When BD Horizon Brilliant Stain Buffer is used in in the multicolor panel, it should also be used in the corresponding compensation controls for all dyes to achieve the most accurate compensation. For the most accurate compensation, compensation controls created with either cells or beads should be exposed to BD Horizon Brilliant Stain Buffer for the same length of time as the corresponding multicolor panel. More information can be found in the Technical Data Sheet of the BD Horizon Brilliant Stain Buffer (Cat. No. 563794/566349) or the BD Horizon Brilliant Stain Buffer Plus (Cat. No. 566385).
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.
- Source of all serum proteins is from USDA inspected abattoirs located in the United States.
- Alexa Fluor® is a registered trademark of Molecular Probes, Inc., Eugene, OR.
- For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
- Cy is a trademark of GE Healthcare.
- BD Horizon Brilliant Violet 711 is covered by one or more of the following US patents: 8,110,673; 8,158,444; 8,227,187; 8,455,613; 8,575,303; 8,354,239.
- BD Horizon Brilliant Stain Buffer is covered by one or more of the following US patents: 8,110,673; 8,158,444; 8,575,303; 8,354,239.
- Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
Companion Products
Stain Buffer (FBS) RUO
Size
500 ML
Cat No.
554656
Stain Buffer (BSA) RUO
Size
500 ML
Cat No.
554657
Fixation and Permeabilization Solution RUO
Size
125 ML
Cat No.
554722
Perm/Wash Buffer RUO
Size
100 ML
Cat No.
554723
Protein Transport Inhibitor (Containing Monensin) RUO
Size
0.7 ML
Cat No.
554724
BV711 Rat IgG2b, κ Isotype Control RUO
Size
50 µg
Cat No.
563045
564081 Rev. 2
Antibody Details
JES5-16E3
The JES5-16E3 monoclonal antibody specifically binds to the mouse cytokine, Interleukin-10 (IL-10). IL-10 is also known as Cytokine Synthesis Inhibitory Factor (CSIF). It is produced by various activated cell types including CD4+ T cells, CD8+ T cells, T regulatory cells, NK T cells, B1 B cells, NK cells, macrophages, dendritic cells, mast cells, granulocytes and keratinocytes. IL-10 plays a pivotal role in regulating immune responses and protecting the host from damage caused by inflammatory and autoimmune responses. IL-10 has numerous biological activities including the inhibition of cytokine synthesis by activated T cells, NK cells, monocytes, and macrophages. In the presence of accessory cells, IL-10 inhibits mitogen- or anti-CD3 induced proliferation of T lymphocytes. IL-10 has also been shown to costimulate the development of thymocytes, B cell differentiation and the generation of cytotoxic T cells. The immunogen used to generate the JES5-16E3 hybridoma was recombinant mouse IL-10. JES5-16E3 is a neutralizing antibody.
564081 Rev. 2
Format Details
BV711
The BD Horizon Brilliant Violet™ 711 (BV711) Dye is part of the BD Horizon Brilliant Violet™ family of dyes. This tandem fluorochrome is comprised of a BV421 donor with an excitation maximum (Ex Max) of 407-nm and an acceptor dye with an emission maximum (Em Max) at 713-nm. BV711, driven by BD innovation, is designed to be excited by the violet laser (405-nm) and detected using an optical filter centered near 710-nm (e.g., a 712/20-nm bandpass filter). The acceptor dye can be excited by the Red (628–640-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.
BV711
407 nm
713 nm
564081 Rev.2
Citations & References
View product citations for antibody "564081" on CiteAb
Development References (4)
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Andersson U, Andersson J. Immunolabeling of cytokine-producing cells in tissues and in suspension. In: Fradelizie D, Emelie D, ed. Cytokine Producing Cells. Paris: Inserm; 1994:32-49.
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Litton MJ, Sander B, Murphy E, O'Garra A, Abrams JS. Early expression of cytokines in lymph nodes after treatment in vivo with Staphylococcus enterotoxin B. J Immunol Methods. 1994; 175(1):47-58. (Clone-specific: Neutralization). 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: Flow cytometry). View Reference
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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: Immunocytochemistry (cytospins), Neutralization). View Reference
564081 Rev. 2
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For Research Use Only. Not for use in diagnostic or therapeutic procedures.