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BV421 Rat Anti-Mouse CD25
BV421 Rat Anti-Mouse CD25

Flow cytometric analysis of CD25 expression on unstimulated and stimulated mouse splenocytes.

       Left and Middle Panels: Mouse splenic leucocytes were preincubated with Purified Rat Anti-Mouse CD16/CD32 antibody (Mouse BD Fc Block™) (Cat No. 553141/553142). The cells were then stained with APC Rat Anti-Mouse CD4 antibody (Cat. No. 553051/561091) and either BD Horizon™ BV421 Rat IgM, κ Isotype Control (Cat. No. 562708; Left Panel) or BD Horizon BV421 Rat Anti-Mouse CD25 antibody (Cat. No. 564571; Middle Panel). Two-color flow cytometric dot plots show the correlated expression patterns for CD25 (or Ig Isotype control staining) versus CD4 for events with the forward and side light-scatter characteristics of viable splenocytes.

       Right Panel: Mouse splenic leucocytes were stimulated for 3 days with concanavalin A. The cells were preincubated with Purified Rat Anti-Mouse CD16/CD32 antibody and then stained with either BD Horizon BV421 Rat IgM, κ Isotype Control (dashed line histogram) or BD Horizon BV421 Rat Anti-Mouse CD25 antibody (solid line histogram). The fluorescence histograms were derived from events with the forward and side light-scatter characteristics of viable lymphoblasts.

       Flow cytometric analysis was performed using a BD™ LSR II Flow Cytometer System.

Flow cytometric analysis of CD25 expression on unstimulated and stimulated mouse splenocytes.

       Left and Middle Panels: Mouse splenic leucocytes were preincubated with Purified Rat Anti-Mouse CD16/CD32 antibody (Mouse BD Fc Block™) (Cat No. 553141/553142). The cells were then stained with APC Rat Anti-Mouse CD4 antibody (Cat. No. 553051/561091) and either BD Horizon™ BV421 Rat IgM, κ Isotype Control (Cat. No. 562708; Left Panel) or BD Horizon BV421 Rat Anti-Mouse CD25 antibody (Cat. No. 564571; Middle Panel). Two-color flow cytometric dot plots show the correlated expression patterns for CD25 (or Ig Isotype control staining) versus CD4 for events with the forward and side light-scatter characteristics of viable splenocytes.

       Right Panel: Mouse splenic leucocytes were stimulated for 3 days with concanavalin A. The cells were preincubated with Purified Rat Anti-Mouse CD16/CD32 antibody and then stained with either BD Horizon BV421 Rat IgM, κ Isotype Control (dashed line histogram) or BD Horizon BV421 Rat Anti-Mouse CD25 antibody (solid line histogram). The fluorescence histograms were derived from events with the forward and side light-scatter characteristics of viable lymphoblasts.

       Flow cytometric analysis was performed using a BD™ LSR II Flow Cytometer System.

Product Details
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BD Horizon™
IL-2 Receptor α chain, p55
Mouse (QC Testing)
Rat LEW, also known as Lewis IgM, κ
IL-2-dependent BALB/c mouse helper T-cell clone HT-2
Flow cytometry (Routinely Tested)
0.2 mg/ml
AB_2738849
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 BV421 under optimum conditions, and unconjugated antibody and free BD Horizon BV421 were removed.

Product Notices

  1. Since applications vary, each investigator should titrate the reagent to obtain optimal results.
  2. An isotype control should be used at the same concentration as the antibody of interest.
  3. 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.
  4. Source of all serum proteins is from USDA inspected abattoirs located in the United States.
  5. Pacific Blue™ is a trademark of Molecular Probes, Inc., Eugene, OR.
  6. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
  7. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
564571 Rev. 1
Antibody Details
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7D4

The 7D4 monoclonal antibody specifically binds to CD25, the low affinity IL-2 Receptor (IL-2Rα, p55) expressed on activated T and B lymphocytes from all mouse strains tested. IL-2Rα by itself is not a signaling receptor. However, it can combine with IL-2 Receptor β (CD122) and γ (CD132) chains to form high-affinity, signaling receptor complexes for IL-2. Resting T and B lymphocytes as well as resting and activated NK cells do not express IL-2Rα.  CD25 is transiently expressed at a low level during normal B-cell development in the bone marrow on the CD45R/B220low TdT- sIg- Pre-B/Pre-B-II and CD45R/B220low TdT- sIgM+ sIgD- immature B stages, but not on the CD45R/B220low TdT+ sIg- Pro-B/Pre-B-I stage nor on CD45R/B220high TdTsIgM+ sIgD+ mature B cells.  It is expressed at a higher level during a very early stage of T-cell development in fetal and adult thymus. Peripheral CD25+ CD4+ T lymphocytes, called regulatory T (Treg) cells, are involved in the maintenance of self-tolerance. It has also been reported that dendritic cells express CD25, which is recognized by mAb 7D4. The 3C7 antibody recognizes an epitope of CD25 which is distinct from those recognized by mAbs 7D4 and PC61, and it blocks binding of IL-2 to CD25.

The antibody was conjugated to BD Horizon BV421 which is part of the BD Horizon Brilliant™ Violet family of dyes. With an Ex Max of 407-nm and Em Max at 421-nm, BD Horizon BV421 can be excited by the violet laser and detected in the standard Pacific Blue™ filter set (eg, 450/50-nm filter). BD Horizon BV421 conjugates are very bright, often exhibiting a 10 fold improvement in brightness compared to Pacific Blue conjugates.

564571 Rev. 1
Format Details
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BV421
The BD Horizon Brilliant Violet™ 421 (BV421) Dye is part of the BD Horizon Brilliant Violet™ family of dyes. This polymer-technology based dye has an excitation maximum (Ex Max) of 407-nm and an emission maximum (Em Max) at 423-nm. Driven by BD innovation, BV421 is designed to be excited by the violet laser (405-nm) and detected using an optical filter centered near 420-nm (e.g., a 431/28-nm or 450/50-nm bandpass filter). BV421 is an ideal alternative for V450 as it is approximately ten times brighter with less spillover into the BV510/V500 detector. Please ensure that your instrument’s configurations (lasers and optical filters) are appropriate for this dye.
altImg
BV421
Violet 405 nm
407 nm
423 nm
564571 Rev.1
Citations & References
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Development References (14)

  1. Chen J, Ma A, Young F, Alt FW. IL-2 receptor alpha chain expression during early B lymphocyte differentiation. Int Immunol. 1994; 6(8):1265-1268. (Biology). View Reference
  2. Crowley M, Inaba K, Witmer-Pack M, Steinman RM. The cell surface of mouse dendritic cells: FACS analyses of dendritic cells from different tissues including thymus. Cell Immunol. 1989; 118(1):108-125. (Clone-specific: Cytotoxicity). View Reference
  3. Garni-Wagner BA, Witte PL, Tutt MM, et al. Natural killer cells in the thymus. Studies in mice with severe combined immune deficiency. J Immunol. 1990; 144(3):796-803. (Clone-specific: Flow cytometry, Fluorescence activated cell sorting). View Reference
  4. Godfrey DI, Kennedy J, Mombaerts P, Tonegawa S, Zlotnik A. Onset of TCR-β gene rearrangement and role of TCR-β expression during CD3-CD4-CD8- thymocyte differentiation. J Immunol. 1994; 152(10):4783-4792. (Clone-specific: Flow cytometry, Fluorescence activated cell sorting). View Reference
  5. Habu S, Okumura K, Diamantstein T, Shevach EM. Expression of interleukin 2 receptor on murine fetal thymocytes. Eur J Immunol. 1985; 15(5):456-460. (Clone-specific: Flow cytometry). View Reference
  6. Malek TR, Robb RJ, Shevach EM. Identification and initial characterization of a rat monoclonal antibody reactive with the murine interleukin 2 receptor-ligand complex. Proc Natl Acad Sci U S A. 1983; 80(18):5694-5698. (Immunogen: Flow cytometry, Functional assay, Immunoprecipitation, Inhibition). View Reference
  7. Malek TR, Schmidt JA, Shevach EM. The murine IL 2 receptor. III. Cellular requirements for the induction of IL 2 receptor expression on T cell subpopulations. J Immunol. 1985; 134(4):2405-2413. (Clone-specific: Flow cytometry). View Reference
  8. Moreau JL, Nabholz M, Diamantstein T, Malek T, Shevach E, Theze J. Monoclonal antibodies identify three epitope clusters on the mouse p55 subunit of the interleukin 2 receptor: relationship to the interleukin 2-binding site. Eur J Immunol. 1987; 17(7):929-935. (Clone-specific: Blocking, Flow cytometry). View Reference
  9. Ortega G, Robb RJ, Shevach EM, Malek TR. The murine IL 2 receptor. I. Monoclonal antibodies that define distinct functional epitopes on activated T cells and react with activated B cells. J Immunol. 1984; 133(4):1970-1975. (Clone-specific: Flow cytometry). View Reference
  10. Pollard AM, Lipscomb MF. Characterization of murine lung dendritic cells: similarities to Langerhans cells and thymic dendritic cells. J Exp Med. 1990; 172(1):159-167. (Clone-specific: Depletion, Fluorescence microscopy, Immunofluorescence). View Reference
  11. Read S, Malmstrom V, Powrie F. Cytotoxic T lymphocyte-associated antigen 4 plays an essential role in the function of CD25(+)CD4(+) regulatory cells that control intestinal inflammation. J Exp Med. 2000; 192(2):295-302. (Clone-specific: Flow cytometry, Fluorescence activated cell sorting). View Reference
  12. Rolink A, Grawunder U, Winkler TH, Karasuyama H, Melchers F. IL-2 receptor alpha chain (CD25, TAC) expression defines a crucial stage in pre-B cell development. Int Immunol. 1994; 6(8):1257-1264. (Biology). View Reference
  13. Takahashi T, Tagami T, Yamazaki S, et al. Immunologic self-tolerance maintained by CD25(+)CD4(+) regulatory T cells constitutively expressing cytotoxic T lymphocyte-associated antigen 4. J Exp Med. 2000; 192(2):303-309. (Clone-specific: Flow cytometry, Fluorescence activated cell sorting). View Reference
  14. Taniguchi T, Minami Y. The IL-2/IL-2 receptor system: a current overview. Cell. 1993; 73(1):5-8. (Biology). View Reference
View All (14) View Less
564571 Rev. 1

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For Research Use Only. Not for use in diagnostic or therapeutic procedures.