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Multicolor flow cytometric analysis of CD25 expression on unstimulated Human peripheral blood lymphocytes. Human whole blood was stained with BD Horizon™ BUV395 Mouse Anti-Human CD4 antibody (Cat. No. 564724) and either BD Horizon™ PE-CF594 Mouse IgG1, κ Isotype Control (Cat. No. 562292; Left Plot) or BD Horizon™ PE-CF594 Mouse Anti-Human CD25 antibody (Cat. No. 567489/567490; Right Plot). Erythrocytes were lysed with BD FACS™ Lysing Solution (Cat. No. 349202). The bivariate pseudocolor density plots showing the correlated expression of CD25 (or Ig Isotype control staining) versus CD4 were derived from gated events with the forward and side light-scatter characteristics of intact lymphocytes. Flow cytometry and data analysis were performed using a BD LSRFortessa™ X-20 Cell Analyzer System and FlowJo™ software.
CD25 expression on activated lymphocytes. Human peripheral blood mononuclear cells (PBMC) were stimulated with PHA (3 days). Cells were stained with BD Horizon™ PE-CF594 Mouse IgG1, κ Isotype Control (dashed line histogram) or BD Horizon™ PE-CF594 Mouse Anti-Human CD25 antibody (solid line histogram). Fluorescent histograms showing CD25 expression (or Ig Isotype control staining) histogram was derived from gated events with light-scatter characteristics of viable (DAPI-negative) lymphoblasts. BD LSRFortessa™ X-20 Cell Analyzer System and FlowJo™ software were used.
BD Horizon™ PE-CF594 Mouse Anti-Human CD25
BD Horizon™ PE-CF594 Mouse Anti-Human CD25
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Preparation And Storage
Recommended Assay Procedures
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 cells and BD® CompBeads to ensure that BD® CompBeads are appropriate for your specific cellular application.
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Companion Products
The BC96 monoclonal antibody specifically binds to human CD25, the low-affinity alpha subunit of the Interleukin-2 Receptor (IL-2Rα), which is also known as TAC antigen. IL2RA encodes CD25 which is a 55 kDa type I transmembrane glycoprotein comprised of an extracellular region with two Complement Control Protein domains (CCP) followed by a transmembrane region and a short cytoplasmic tail. CD25 is constitutively expressed at high levels on natural T regulatory cells and variably expressed on conventional T cells and B cells and their precursors, NK cells, monocytes, and macrophages. CD25 expression can be highly upregulated upon antigenic or mitogenic stimulation of T cells or B cells. A soluble form of CD25 is found in biological fluids due to proteolytic cleavage of the extracellular region of transmembrane CD25. CD25 noncovalently associates with CD122 (IL-2Rβ chain) and CD132 (IL-2Rγ, also known as the common γ chain or γc) to form the high-affinity signal-transducing IL-2R complex (IL-2Rαβγ). This heterotrimeric receptor mediates biological activities of IL-2 which can act as a cellular activation, growth, and differentiation factor and regulator of cell viability. Analysis of CD25 expression can be used to characterize the nature of normal leucocytes in their resting states or activated during inflammatory or immune responses as well as those present in certain disease states.
Development References (4)
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Chapel A, Bensussan A, Vilmer E, Dormont D. Differential human immunodeficiency virus expression in CD4+ cloned lymphocytes: from viral latency to replication.. J Virol. 1992; 66(6):3966-70. (Immunogen: Flow cytometry). View Reference
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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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Poszepczynska E, Bagot M, Echchakir H, et al. Functional characterization of an IL-7-dependent CD4(+)CD8alphaalpha(+) Th3-type malignant cell line derived from a patient with a cutaneous T-cell lymphoma.. 2000; 96(3):1056-63. (Clone-specific). View Reference
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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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For Research Use Only. Not for use in diagnostic or therapeutic procedures.
Refer to manufacturer's instructions for use and related User Manuals and Technical Data Sheets before using this product 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.