BD Horizon™ RY743 Hamster Anti-Helios
Clone 22F6 (RUO)
Two-color flow cytometric analysis of Helios expression Left Panel: Human peripheral blood mononuclear cells (PBMCs) were stained with BD Horizon™ BV421 Mouse Anti-Human CD4 antibody (Cat. No. 566907) and fixed and permeabilized with the BD Pharmingen™ Transcription Factor Buffer Set (Cat. No. 562574/562725). The cells were then stained with either BD Horizon™ RY743 Hamster IgG2, κ Isotype Control (Cat. No. 572208; Left Plot; used as a representative Hamster IgG Isotype Control) or BD Horizon™ RY743 Hamster Anti-Helios antibody (Cat. No. 572206/572207; Right Plot). Right Panel: Mouse splenic leukocytes were stained with BD Horizon™ BV421 Rat Anti-Mouse CD4 antibody (Cat. No. 562891) and fixed and permeabilized with the BD Pharmingen™ Transcription Factor Buffer Set. The cells were then stained with either BD Horizon™ RY743 Hamster IgG2, κ Isotype Control (Left Plot) or BD Horizon™ RY743 Hamster Anti-Helios antibody (Right Plot). The bivariate pseudocolor density plots show the correlated expression of Helios (or Ig Isotype control staining) versus CD4 were derived from gated events with the forward and side light-scatter characteristics of intact Human peripheral blood lymphocytes or Mouse splenic leukocytes. Flow cytometry and data analysis were performed using a BD FACSymphony™ A5 SE Cell Analyzer System and FlowJo™ Software.
Two-color flow cytometric analysis of Helios expression Left Panel: Human peripheral blood mononuclear cells (PBMCs) were stained with BD Horizon™ BV421 Mouse Anti-Human CD4 antibody (Cat. No. 566907) and fixed and permeabilized with the BD Pharmingen™ Transcription Factor Buffer Set (Cat. No. 562574/562725). The cells were then stained with either BD Horizon™ RY743 Hamster IgG2, κ Isotype Control (Cat. No. 572208; Left Plot; used as a representative Hamster IgG Isotype Control) or BD Horizon™ RY743 Hamster Anti-Helios antibody (Cat. No. 572206/572207; Right Plot). Right Panel: Mouse splenic leukocytes were stained with BD Horizon™ BV421 Rat Anti-Mouse CD4 antibody (Cat. No. 562891) and fixed and permeabilized with the BD Pharmingen™ Transcription Factor Buffer Set. The cells were then stained with either BD Horizon™ RY743 Hamster IgG2, κ Isotype Control (Left Plot) or BD Horizon™ RY743 Hamster Anti-Helios antibody (Right Plot). The bivariate pseudocolor density plots show the correlated expression of Helios (or Ig Isotype control staining) versus CD4 were derived from gated events with the forward and side light-scatter characteristics of intact Human peripheral blood lymphocytes or Mouse splenic leukocytes. Flow cytometry and data analysis were performed using a BD FACSymphony™ A5 SE Cell Analyzer System and FlowJo™ Software.
Product Details
BD Horizon™
IKAROS family zinc finger protein 2; ANF1A2; HELIOS; ZNF1A2; ZNFN1A2
Human (QC Testing), Mouse (Tested in Development)
Armenian Hamster IgG
Mouse Helios Peptide
Intracellular staining (flow cytometry) (Routinely Tested)
5 µl/test
Aqueous buffered solution containing ≤0.09% sodium azide.
RUO
Preparation And Storage
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 unreacted dye was removed. Store undiluted at 4°C and protected from prolonged exposure to light. Do not freeze.
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.
Product Notices
- Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
- An isotype control should be used at the same concentration as the antibody of interest.
- Species cross-reactivity detected in product development may not have been confirmed on every format and/or application.
- When using high concentrations of antibody, background binding of this dye to erythroid fragments produced by ammonium chloride-based lysis, such as with BD Pharm Lyse™ Lysing Buffer (Cat. No. 555899), has been observed when the antibody conjugate was present during the lysis procedure. This may cause nonspecific staining of target cells, such as leukocytes, which have bound the resulting erythroid fragments. This background can be mitigated by any of the following: titrating the antibody conjugate to a lower concentration, fixing samples with formaldehyde, or removing erythrocytes before staining (eg, gradient centrifugation or pre-lysis with wash). This background has not been observed when cells were lysed with BD FACS™ Lysing Solution (Cat. No. 349202) after staining.
- 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.
- 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).
- For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
- Please refer to http://regdocs.bd.com to access safety data sheets (SDS).
- Human donor specific background has been observed in relation to the presence of anti-polyethylene glycol (PEG) antibodies, developed as a result of certain vaccines containing PEG, including some COVID-19 vaccines. We recommend use of BD Horizon Brilliant™ Stain Buffer in your experiments to help mitigate potential background. For more information visit https://www.bdbiosciences.com/en-us/support/product-notices.
- Please observe the following precautions: We recommend that special precautions be taken (such as wrapping vials, tubes, or racks in aluminum foil) to protect exposure of conjugated reagents, including cells stained with those reagents, to any room illumination. Absorption of visible light can significantly affect the emission spectra and quantum yield of tandem fluorochrome conjugates.
- Tandem fluorochromes contain both an energy donor and an energy acceptor. Although every effort is made to minimize the lot-to-lot variation in the efficiency of the fluorochrome energy transfer, differences in the residual emission from the donor may be observed. Additionally, multi-laser cytometers may directly excite both the donor and acceptor fluorochromes. Therefore, we recommend for every tandem conjugate, a matched individual single-stain control be acquired for generating a compensation or spectral unmixing matrix.
- Cy is a trademark of Global Life Sciences Solutions Germany GmbH or an affiliate doing business as Cytiva.
- For U.S. patents that may apply, see bd.com/patents.
Companion Products
Transcription Factor Buffer Set RUO
Size
100 Tests
Cat No.
562574
RY743 Hamster IgG2, κ Isotype Control RUO
Size
0.1 mg
Cat No.
572208
572207 Rev. 1
Antibody Details
22F6
The 22F6 monoclonal antibody specifically binds to mouse and human Helios. Helios is a member of the Ikaros family of zinc-finger transcription factors, which play important roles in hematopoietic cell development and tumor suppression. Helios expression is restricted to the earliest stages of embryonic hematopoiesis, a variety of epithelial tissues and is notably increased in thymic-derived regulatory CD4+Foxp3+ T (Treg) cells. Its high expression levels in Treg cells are independent from Foxp3 and are believed to contribute, along with other transcription factors, to the phenotypic stability of natural regulatory T cells. Accordingly, it has been demonstrated that Helios directly stimulates Foxp3 transcription while it inhibits ll2 gene expression, contributing for the maintenance of cellular anergy. Helios is also differentially expressed during negative and positive selection in the thymus, marking CD4+ autoreactive cells for deletion. Helios may possibly play roles in T cell activation, since it is upregulated in Th2 and Tfh cells. Despite these roles in T cell development and function, Helios genetic ablation in mice revealed no significant abnormalities in Treg or other T cell subsets. This finding suggests that other Ikaros family members may play redundant roles.
572207 Rev. 1
Format Details
RY743
The BD Horizon RealYellow™ 743 (RY743) Dye is part of the BD® family of yellow-green dyes. It is a tandem fluorochrome with an excitation maximum (Ex Max) at 557 nm and an emission maximum (Em Max) at 743 nm as measured using an antibody-dye conjugate. Driven by BD® innovation, RY743 is optimized for detection on spectral cytometers and designed to be excited by the Yellow-Green laser (561 nm) with minimal excitation by the Blue laser (488 nm). Given its unique emission max, RY743 can be used on a spectral instrument in combination with RY703 and either RY775 or PE-Cy7 to provide an additional color excited by the Yellow-Green laser. For conventional instruments equipped with a Yellow-Green laser, we recommend using an optical filter centered near 750 nm (e.g., a 750/60-nm bandpass filter). Please ensure that your instrument configuration (lasers and optical filters) is appropriate for this dye.
RY743
Yellow-Green 561 nm
557 nm
743 nm
572207 Rev.1
Citations & References
View product citations for antibody "572207" on CiteAb
Development References (10)
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Baine I, Basu S, Ames R, Sellers RS, Macian F. Helios induces epigenetic silencing of IL2 gene expression in regulatory T cells. J Immunol. 2013; 190(3):1008-1016. (Biology). View Reference
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Cai Q, Dierich A, Oulad-Abdelghani M, Chan S, Kastner P. Helios deficiency has minimal impact on T cell development and function. J Immunol. 183(4)(Biology). View Reference
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Daley SR, Hu DY, Goodnow CC. Helios marks strongly autoreactive CD4+ T cells in two major waves of thymic deletion distinguished by induction of PD-1 or NF-kappaB. J Exp Med. 2013; 210(2):269-285. (Biology). View Reference
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Fu W, Ergun A, Lu T, et al. A multiply redundant genetic switch 'locks in' the transcriptional signature of regulatory T cells. Nat Immunol. 2012; 13(10):972-980. (Biology). View Reference
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Georgopoulos K. Haematopoietic cell-fate decisions, chromatin regulation and ikaros. Nat Rev Immunol. 2002; 2(3):162-174. (Biology). View Reference
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Kelley CM, Ikeda T, Koipally J, et al. Helios, a novel dimerization partner of Ikaros expressed in the earliest hematopoietic progenitors. Curr Biol. 1998; 8(9):508-515. (Biology). View Reference
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Serre K, Benezech C, Desanti G, et al. Helios is associated with CD4 T cells differentiating to T helper 2 and follicular helper T cells in vivo independently of Foxp3 expression. PLoS ONE. 2011; 6(6):e20731. (Biology). View Reference
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Sugimoto N, Oida T, Hirota K, et al. Foxp3-dependent and -independent molecules specific for CD25+CD4+ natural regulatory T cells revealed by DNA microarray analysis. Int Immunol. 18(8)(Biology). View Reference
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Thornton AM, Korty PE, Tran DQ, et al. Expression of Helios, an Ikaros transcription factor family member, differentiates thymic-derived from peripherally induced Foxp3+ T regulatory cells. J Immunol. 2010; 184(7):3433-3441. (Immunogen: ELISA, Flow cytometry, Western blot). View Reference
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Yoshimatsu Y, Sujino T, Miyamoto K, et al. Aryl hydrocarbon receptor signals in epithelial cells govern the recruitment and location of Helios+ Tregs in the gut.. Cell Rep. 2022; 39(6):110773. (Clone-specific: Flow cytometry). View Reference
572207 Rev. 1
Please refer to Support Documents for Quality Certificates
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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.