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BV605 Mouse Anti-Human CD69
BV605 Mouse Anti-Human CD69

Flow cytometric analysis of CD69 expressed on stimulated peripheral blood mononuclear cells. Human PBMC were stimulated for 24 hours with phytohemagglutinin. Cells were then stained with either a BD Horizon™ BV605 Mouse IgG1, κ Isotype control (Cat. No. 562652; dashed line histogram) or with the BD Horizon™ BV605 Mouse Anti-Human CD69 antibody (Cat. No. 562989; solid line histogram). The fluorescence histograms were derived from gated events with the forward and side light-scatter characteristics of viable activated lymphocytes. Flow cytometry was performed using a BD™ LSR II Flow Cytometer System.

Flow cytometric analysis of CD69 expressed on stimulated peripheral blood mononuclear cells. Human PBMC were stimulated for 24 hours with phytohemagglutinin. Cells were then stained with either a BD Horizon™ BV605 Mouse IgG1, κ Isotype control (Cat. No. 562652; dashed line histogram) or with the BD Horizon™ BV605 Mouse Anti-Human CD69 antibody (Cat. No. 562989; solid line histogram). The fluorescence histograms were derived from gated events with the forward and side light-scatter characteristics of viable activated lymphocytes. Flow cytometry was performed using a BD™ LSR II Flow Cytometer System.

Product Details
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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)
969
AB_2737935
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™ BV605 under optimum conditions, and unconjugated antibody and free BD Horizon™ BV605 were removed.

Recommended Assay Procedures

For optimal and reproducible results, BD Horizon Brilliant Stain Buffer should be used anytime two or more BD Horizon Brilliant dyes 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).

Product Notices

  1. 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).
  2. An isotype control should be used at the same concentration as the antibody of interest.
  3. Source of all serum proteins is from USDA inspected abattoirs located in the United States.
  4. 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.
  5. 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.
  6. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
  7. 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 BD Horizon™ BV421 may be observed. Therefore, we recommend that individual compensation controls be performed for every BD Horizon™ BV605 conjugate.
  8. CF™ is a trademark of Biotium, Inc.
  9. BD Horizon Brilliant Violet 605 is covered by one or more of the following US patents: 8,110,673; 8,158,444; 8,227,187; 8,455,613; 8,575,303; 8,354,239.
  10. 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.
  11. Species cross-reactivity detected in product development may not have been confirmed on every format and/or application.
  12. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
562989 Rev. 3
Antibody Details
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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.

Clone FN50 reacts with the human form of the 28/34 kDa dimeric glycoprotein expressed early during activation of lymphocytes, monocytes, and platelets. It also cross-reacts with a subset of peripheral blood mononuclear cells (lymphocytes and monocytes) of rhesus and cynomolgus macaque monkeys. The distribution on lymphocytes is similar to that observed with human peripheral blood lymphocytes with the majority of the cells demonstrating an increase in FN50 positivity following overnight incubation with phorbol myristrate acetate (PMA).

This antibody is conjugated to BD Horizon BV605 which is part of the BD Horizon Brilliant™ Violet family of dyes. With an Ex Max of 407-nm and Em Max of 602-nm, BD Horizon BV605 can be excited by a violet laser and detected with a standard 610/20-nm filter set. BD Horizon BV605 is a tandem fluorochrome of BD Horizon BV421 and an acceptor dye with an Em max at 605-nm. Due to the excitation of the acceptor dye by the green (532 nm) and yellow-green (561 nm) lasers, there will be significant spillover into the PE and BD Horizon PE-CF594 detectors off the green or yellow-green lasers. BD Horizon BV605 conjugates are very bright, often exhibiting brightness equivalent to PE conjugates and can be used as a third color off of the violet laser.

562989 Rev. 3
Format Details
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BV605
The BD Horizon Brilliant Violet™ 605 (BV605) dye is part of the BD Horizon Brilliant Violet™ family of dyes. This tandem fluorochrome is comprised of a BV421 donor with an excitation maximum (Ex Max) of 407-nm and an acceptor dye with an emission maximum (Em Max) at 605-nm. BV605, driven by BD innovation, is designed to be excited by the violet laser (405-nm) and detected using an optical filter centered near 610-nm (e.g., a 610/20-nm bandpass filter). The acceptor dye can be excited by the yellow-green (561-nm) laser resulting in cross-laser excitation and fluorescence spillover. Please ensure that your instrument’s configurations (lasers and optical filters) are appropriate for this dye.
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BV605
Violet 405 nm
407 nm
605 nm
562989 Rev.3
Citations & References
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Development References (3)

  1. CD69. In: Zola H. Leukocyte and stromal cell molecules : the CD markers. Hoboken, N.J.: Wiley-Liss; 2007:161.
  2. Knapp W. W. Knapp .. et al., ed. Leucocyte typing IV : white cell differentiation antigens. Oxford New York: Oxford University Press; 1989:1-1182.
  3. 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.
562989 Rev. 3

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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.