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Two-color flow cytometric analysis of CD25 expression on unstimulated human lymphocytes. Whole blood was stained with APC Mouse Anti-Human CD4 antibody (561841/555349/561840) and either BD Horizon™ BV480 Mouse IgG1, κ Isotype Control (Cat. No. 565652; Left Plot) or BD Horizon BV480 Mouse Anti-Human CD25 antibody (Cat. No. 566102/566160; Right Plot). Erythrocytes were lysed with BD FACS Lysing Solution (Cat. No. 349202). The two-color flow cytometric contour plot showing the correlated expression of CD25 (or Ig Isotype control staining) versus CD4 was derived from gated events with the forward and side light-scatter characteristics of intact lymphocytes. Flow cytometric analysis was performed using a BD LSRFortessa™ Cell Analyzer System.

Flow cytometric analysis of CD25 expression on stimulated human lymphocytes. Phytohemagglutinin (PHA)-stimulated (3 days) peripheral blood mononuclear cells were stained with either BD Horizon™ BV480 Mouse IgG1, κ Isotype Control (dashed line histogram) or BD Horizon BV480 Mouse Anti-Human CD25 antibody (solid line histogram). The fluorescence histogram showing CD25 expression (or Ig Isotype control staining) was derived from gated events with the forward and side light-scatter characteristics of viable lymphoblasts. Flow cytometric analysis was performed using a BD LSRFortessa™ Cell Analyzer System.


BD Horizon™ BV480 Mouse Anti-Human CD25

BD Horizon™ BV480 Mouse Anti-Human CD25

Regulatory Status Legend
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
Recommended Assay Procedures
For optimal and reproducible results, BD Horizon Brilliant Stain Buffer should be used anytime two or more BD Horizon Brilliant dyes (including BD Optibuild Brilliant reagents) are used in the same experiment. Fluorescent dye interactions may cause staining artifacts which may affect data interpretation. The BD Horizon Brilliant Stain Buffer was designed to minimize these interactions. 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).
For Immunofluorescence Applications:
The use of a mounting reagent (eg, ProLong® Gold) is highly recommended to maximize the photostability of BV480. For confocal microscopy systems, a 440 nm laser is the optimal excitation source and the recommended emission filter is a 485/20 nm bandpass filter.
For epifluorescence microscopes with broad spectrum excitation sources, the recommended excitation and emission filters are 445/20 nm and 485/20 nm bandpass filters, respectively. For specific multicolor imaging applications, the exact filter configurations should be optimized by the end user. For additional instrument/filter configuration information, please visit http://www.bdbiosciences.com/research/cellularimaging.
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.
- Source of all serum proteins is from USDA inspected abattoirs located in the United States.
- 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.
- BD Horizon Brilliant Violet 480 is covered by one or more of the following US patents: 8,575,303; 8,354,239.
- Species cross-reactivity detected in product development may not have been confirmed on every format and/or application.
- 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.
- BD Horizon Brilliant Violet 480 is covered by one or more of the following US patents: 8,575,303; 8,354,239.
- ProLong® is a registered trademark of Thermo Fisher Scientific, Inc. Waltham, MA.
- Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
Companion Products






The M-A251 monoclonal antibody specifically binds to the 55 kDa type I transmembrane glycoprotein known as low-affinity interleukin-2 receptor alpha chain subunit (IL-2Rα). CD25 is expressed on regulatory T cells, activated lymphocytes (T and B), and monocytes. It associates with the IL-2Rβ/CD122 and IL-2Rγ/CD132 receptor chains to form the high-affinity IL-2R complex. CD25 expression on T and B lymphocytes is upregulated by antigenic or mitogenic stimulation. Soluble CD25/IL-2Rα is produced as a consequence of lymphocyte stimulation and is found in biological fluids following inflammatory responses.
The antibody was conjugated to BD Horizon BV480 which is part of the BD Horizon Brilliant™ Violet family of dyes. With an Ex Max of 436-nm and Em Max at 478-nm, BD Horizon BV480 can be excited by the violet laser and detected in the BD Horizon BV510 (525/40-nm) filter set. BV480 has less spillover into the BV605 detector and, in general, is brighter than BV510.

Development References (7)
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Janszen M, Buck D, Maino VC. Functional and molecular properties of CD25 monoclonal antibodies. In: Knapp W. W. Knapp .. et al., ed. Leucocyte typing IV : white cell differentiation antigens. Oxford New York: Oxford University Press; 1989:403-406.
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Knapp W. W. Knapp .. et al., ed. Leucocyte typing IV : white cell differentiation antigens. Oxford New York: Oxford University Press; 1989:1-1182.
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Lin G-X, Yang X, Hollemweguer E, et al. Cross-reactivity of CD antibodies in eight animal species. In: Mason D. David Mason .. et al., ed. Leucocyte typing VII : white cell differentiation antigens : proceedings of the Seventh International Workshop and Conference held in Harrogate, United Kingdom. Oxford: Oxford University Press; 2002:519-523.
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Nakamura K, Kitani A, Fuss I, et al. TGF-β1 plays an important role in the mechanism of CD4+CD25+ regulatory T cell activity in both humans and mice. J Immunol. 2004; 172(2):834-842. (Clone-specific: Flow cytometry, Fluorescence activated cell sorting). View Reference
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Ravoet AM, Latinne D, Couvreur B, et al. CD25 mab: Epitopes recognized, effect on lymphocyte activation, mediation of ADCC. In: Knapp W. W. Knapp .. et al., ed. Leucocyte typing IV : white cell differentiation antigens. Oxford New York: Oxford University Press; 1989:408-411.
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Schlossman SF. Stuart F. Schlossman .. et al., ed. Leucocyte typing V : white cell differentiation antigens : proceedings of the fifth international workshop and conference held in Boston, USA, 3-7 November, 1993. Oxford: Oxford University Press; 1995.
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Schwarting R, Stein H. Cluster report: CD25. In: Knapp W. W. Knapp .. et al., ed. Leucocyte typing IV : white cell differentiation antigens. Oxford New York: Oxford University Press; 1989:399-403.
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.