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BD Horizon™ RB780 Mouse Anti-Human CD69
Clone FN50 (also known as FN 50) (RUO)
Flow cytometric analysis of CD69 expression on stimulated Human peripheral blood mononuclear cells. Human peripheral blood mononuclear cells (PBMC) were stimulated for 24 hours with Phytohemagglutinin (PHA). The cells were then stained with either BD Horizon™ RB780 Mouse IgG1, κ Isotype Control (Cat. No. 568532; dashed line histogram) or BD Horizon™ RB780 Mouse Anti-Human CD69 antibody (Cat. No. 568754/568755; solid line histogram). The fluorescence histogram showing CD69 expression (or Ig Isotype control staining) was derived from gated events with the forward and side light-scatter characteristics of viable activated lymphocytes. Flow cytometry and data analysis were performed using a BD FACSymphony™ A5 SE Cell Analyzer System and FlowJo™ software.
Flow cytometric analysis of CD69 expression on stimulated Human peripheral blood mononuclear cells. Human peripheral blood mononuclear cells (PBMC) were stimulated for 24 hours with Phytohemagglutinin (PHA). The cells were then stained with either BD Horizon™ RB780 Mouse IgG1, κ Isotype Control (Cat. No. 568532; dashed line histogram) or BD Horizon™ RB780 Mouse Anti-Human CD69 antibody (Cat. No. 568754/568755; solid line histogram). The fluorescence histogram showing CD69 expression (or Ig Isotype control staining) was derived from gated events with the forward and side light-scatter characteristics of viable activated lymphocytes. Flow cytometry and data analysis were performed using a BD FACSymphony™ A5 SE Cell Analyzer System and FlowJo™ software.
Product Details
BD Horizon™
AIM; CLEC2C; EA1; GP32/28; Leu23; MLR-3; VEA; BL-AC/P26
Human (QC Testing), Rhesus, Cynomolgus, Baboon (Tested in Development)
Mouse IgG1, κ
Anti-µ stimulated human B lymphocytes
Flow cytometry (Routinely Tested)
5 µl
IV A91 (A091)
AB_3684519
Aqueous buffered solution containing ≤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 to the dye under optimum conditions and unreacted dye was removed.
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
- 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.
- Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
- 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.
- Please observe the following precautions: Absorption of visible light can significantly alter the energy transfer occurring in any tandem fluorochrome conjugate; therefore, we recommend that special precautions be taken (such as wrapping vials, tubes, or racks in aluminum foil) to prevent exposure of conjugated reagents, including cells stained with those reagents, to room illumination.
- 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.
- Species cross-reactivity detected in product development may not have been confirmed on every format and/or application.
- Please refer to http://regdocs.bd.com to access safety data sheets (SDS).
- Cy is a trademark of Global Life Sciences Solutions Germany GmbH or an affiliate doing business as Cytiva.
- 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.
Companion Products
Stain Buffer (FBS) RUO
Size
500 ML
Cat No.
554656
RB780 Mouse IgG1, κ Isotype Control RUO
Size
50 µg
Cat No.
568532
Stain Buffer (BSA) RUO
Size
500 ML
Cat No.
554657
568754 Rev. 2
Antibody Details
FN50
The FN50 monoclonal antibody specifically binds to human CD69. CD69 is also known as activation-induced molecule (AIM), early activation antigen (EA-1), very early activation antigen (VEA), C-type lectin domain family 2 member C (CLEC2C), MLR-3, GP32/28 and Leu-23. CD69 is a transmembrane type II homodimer receptor. CD69 is comprised of disulfide-linked, differentially glycosylated core protein subunits that are approximately 28 and 34 kDa in size. Each subunit contains a C-type lectin domain. CD69 is expressed on activated T, B, and natural killer (NK) lymphocytes, thymocytes, neutrophils, eosinophils and platelets. In normal peripheral blood, a small and variable percentage of lymphocytes typically express detectable membrane CD69 antigen. Upon activation, CD69 antigen expression increases on lymphocytes. Peak CD69 expression generally occurs within 18 hours of activation, preceding the appearance of HLA-DR, IL-2Rα (CD25) and transferrin receptor (CD71). CD69 is highly expressed on the bright CD3+ subset of thymocytes. FN50 monoclonal antibody labels NK cells and most lymphocytes of the follicular mantle and perifollicular/interfollicular zone as well as germinal center T cells of lymph nodes and tonsils. Studies indicate that CD69 serves as a signaling receptor in the activation of a variety of cell types.
568754 Rev. 2
Format Details
RB780
The BD Horizon RealBlue™ 780 (RB780) Dye is part of the BD family of blue dyes. It is a tandem fluorochrome with an excitation maximum (Ex Max) at 498-nm and an emission maximum (Em Max) at 781-nm. Driven by BD innovation, RB780 can be used on both spectral and conventional cytometers and is designed to be excited by the Blue laser (488-nm) with minimal excitation by the 561-nm Yellow-Green laser. For conventional instruments equipped with a Blue laser (488-nm), RB780 can be used as an alternative to PE-Cy7 and we recommend using an optical filter centered near 780-nm (eg, a 780/60-nm bandpass filter). For spectral instruments equipped with a Blue laser (488-nm), it can be used in conjunction with PE-Cy7. RB780 is on average brighter than PE-Cy7 and has minimal spillover into Yellow-Green detectors.
RB780
498 nm
781 nm
568754 Rev.2
Citations & References
View product citations for antibody "568754" on CiteAb
Development References (9)
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Beiske K, AAS-Eng DA, Smeland EB, Sundan A, Marton PF, Funderud S. Immunohistochemical and functional characterization of the mAb A91 (FN 50)-reactive activation antigen (CD69) expressed on subsets of normal and neoplastic lymphoid cells. In: Knapp W. W. Knapp .. et al., ed. Leucocyte typing IV : white cell differentiation antigens. Oxford New York: Oxford University Press; 1989:436-439.
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CD69. In: Zola H. Leukocyte and stromal cell molecules : the CD markers. Hoboken, N.J.: Wiley-Liss; 2007:161.
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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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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, Biedobitek G, Stein H. Cluster report: CD69. In: Knapp W. W. Knapp .. et al., ed. Leucocyte typing IV : white cell differentiation antigens. Oxford New York: Oxford University Press; 1989:428-429.
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Stein H, Schwarting R, Niedobitek G, Dallenbach F. Activation Section Report. In: Knapp W. W. Knapp .. et al., ed. Leucocyte typing IV : white cell differentiation antigens. Oxford New York: Oxford University Press; 1989:387-398.
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Tomescu C, Chehimi J, Maino VC, Montaner LJ. NK cell lysis of HIV-1-infected autologous CD4 primary T cells: requirement for IFN-mediated NK activation by plasmacytoid dendritic cells. 2007; 179(4):2097-2104. (Clone-specific: Flow cytometry). View Reference
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Yoshino N, Ami Y, Terao K, Tashiro F, Honda M. Upgrading of flow cytometric analysis for absolute counts, cytokines and other antigenic molecules of cynomolgus monkeys (Macaca fascicularis) by using anti-human cross-reactive antibodies. Exp Anim. 2000; 49(2):97-110. (Clone-specific: Flow cytometry). View Reference
568754 Rev. 2
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.