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PE-Cy™7 Mouse Anti-Human CD196 (CCR6)
PE-Cy™7 Mouse Anti-Human CD196 (CCR6)

Flow cytometric analysis of CD196 expression on human lysed whole blood. Whole blood was lysed with BD FACS™ Lysing Solution (Cat. No. 349202), then stained with either PE-Cy™7 Mouse Anti-Human CD196 antibody (Cat. No. 560620; empty histogram) or PE-Cy™7Mouse IgG1, κ isotype control (Cat. No. 557872; shaded histogram). Fluorescent histograms depicting CD196 (or Ig isotype control) expression were derived from gated events based on forward and side light scatter characteristics of viable lymphocytes. Flow cytometry was performed on a BD™ LSR II flow cytometry system.

Flow cytometric analysis of CD196 expression on human lysed whole blood. Whole blood was lysed with BD FACS™ Lysing Solution (Cat. No. 349202), then stained with either PE-Cy™7 Mouse Anti-Human CD196 antibody (Cat. No. 560620; empty histogram) or PE-Cy™7Mouse IgG1, κ isotype control (Cat. No. 557872; shaded histogram). Fluorescent histograms depicting CD196 (or Ig isotype control) expression were derived from gated events based on forward and side light scatter characteristics of viable lymphocytes. Flow cytometry was performed on a BD™ LSR II flow cytometry system.

Product Details
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BD Pharmingen™
BN-1; C-C CKR-6; C-C chemokine receptor type 6; CC-CKR-6; CCR-6
Human (QC Testing), Rhesus, Cynomolgus, Baboon (Tested in Development)
Mouse IgG1, κ
Human CD196/CCR6 Peptide
Flow cytometry (Routinely Tested)
5 µl
IX 48
1235
AB_1727440
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 PE-Cy7 under optimum conditions, and unconjugated antibody and free PE-Cy7 were removed.

Recommended Assay Procedures

Flow cytometry:  Chemokine receptors are known to internalize during manipulation resulting in low frequency expression.  Investigators are advised to perform immunophenotyping studies of chemokine receptors on freshly collected samples (<24 Hrs). Incubation with the antibody should be done at 4°C in the dark. Cellular manipulation, such as Ficoll separation, freezing, or exposure to cold temperatures prior to staining should be minimized and have been shown to cause a decrease in staining intensity and/or inconsistent results.

Investigators should note that alternative staining procedures may be neccessary.  A multiple-step staining procedure may be helpful, in some instances, to amplify immunofluorescent signals for the flow cytometric analysis of CD196 expression.  Investigators may find the Purified Mouse Anti-Human CD196 antibody (Cat. No. 559560) to be useful in conjunction with appropriate secondary and tertiary reagents for detecting low frequency expression, such as with Biotin Goat Anti-Mouse Ig (Cat. No. 553999) and PE Streptavidin (Cat. No. 554061) or PE-Cy™7 Streptavidin (Cat. No. 557598).

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. 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.
  5. Warning: Some APC-Cy7 and PE-Cy7 conjugates show changes in their emission spectrum with prolonged exposure to formaldehyde. If you are unable to analyze fixed samples within four hours, we recommend that you use BD™ Stabilizing Fixative (Cat. No. 338036).
  6. PE-Cy7 is a tandem fluorochrome composed of R-phycoerythrin (PE), which is excited by 488-nm light and serves as an energy donor, coupled to the cyanine dye Cy7, which acts as an energy acceptor and fluoresces maximally at 780 nm. PE-Cy7 tandem fluorochrome emission is collected in a detector for fluorescence wavelengths of 750 nm and higher. 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 PE may be observed. Therefore, we recommend that individual compensation controls be performed for every PE-Cy7 conjugate. PE-Cy7 is optimized for use with a single argon ion laser emitting 488-nm light, and there is no significant overlap between PE-Cy7 and FITC emission spectra. When using dual-laser cytometers, which may directly excite both PE and Cy7, we recommend the use of cross-beam compensation during data acquisition or software compensation during data analysis.
  7. 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.
  8. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
  9. Cy is a trademark of GE Healthcare.
  10. Species cross-reactivity detected in product development may not have been confirmed on every format and/or application.
  11. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
560620 Rev. 3
Antibody Details
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11A9

The 11A9 monoclonal antibody specifically binds to CD196, which is also known as CCR6. CCR6 is a seven-transmembrane, G-protein-coupled, glycoprotein receptor that is a member of the beta chemokine receptor family. The human CCR6 gene has been mapped to chromosome 6q27. CCR6 is a receptor for the CC chemokine CCL20/MIP-3alpha/LARC/Exodus and also binds with lower affinity to and mediates responses to beta-defensin2/hBD-2. CCR6 is predominantly expressed by B lymphocytes, certain subsets of effector and memory T cells and by immature dendritic cells but not by monocytes, NK cells, or granulocytes. Skin-homing CLA (Cutaneous Lymphocyte Antigen)-positive memory T cells, Th1 cells, regulatory T cells and IL-17A-producing Th17 cells predominantly express high levels of CCR6. CCR6 mediates the trafficking of T, B, and dendritic cells to epithelial sites near the skin and mucosal surfaces during inflammatory and immunological responses. An N-terminal peptide of human CCR6 was used as an immunogen to generate the 11A9 hybridoma. The 11A9 antibody does not cross-react with human CCR1, CCR2, CCR3, CCR4, CCR5, CCR7, CCR8, CCR9, CXCR1, CXCR2, CXCR3, CXCR4 and CXCR5 receptors. This antibody is NOT a neutralizing antibody.

560620 Rev. 3
Format Details
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PE-Cy7
PE-Cy7 dye is a part of the BD PE family of dyes. This tandem fluorochrome is comprised of a R-Phycoerythrin (PE) donor that has excitation maxima (Ex Max) of 496-nm and 566-nm and an acceptor dye, Cy™7, with an emission maximum (Em Max) at 781-nm. PE can be excited by the Blue (488-nm), Green (532-nm) and yellow-green (561-nm) lasers and detected using an optical filter centered near 781 nm (e.g., a 760/60-nm bandpass filter). The donor dye can be excited by the Blue (488-nm), Green (532-nm) and yellow-green (561-nm) lasers and the acceptor dye can be excited by the Red (627–640-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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PE-Cy7
Yellow-Green 488 nm, 532 nm, 561 nm
496 nm, 566 nm
781 nm
560620 Rev.3
Citations & References
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Development References (10)

  1. Baba M, Imai T, Nishimura M, et al. Identification of CCR6, the specific receptor for a novel lymphocyte-directed CC chemokine LARC. J Biol Chem. 1997; 272(23):14893-14898. (Biology). View Reference
  2. Greaves DR, Wang W, Dairaghi DJ, et al. CCR6, a CC chemokine receptor that interacts with macrophage inflammatory protein 3alpha and is highly expressed in human dendritic cells. J Exp Med. 1997; 186(6):837-844. (Biology). View Reference
  3. Homey B, Dieu-Nosjean MC, Wiesenborn A, et al. Up-regulation of macrophage inflammatory protein-3 alpha/CCL20 and CC chemokine receptor 6 in psoriasis. J Immunol. 2000; 164(12):6621-6632. (Biology). View Reference
  4. Kim CH, Rott L, Kunkel EJ, et al. Rules of chemokine receptor association with T cell polarization in vivo. J Clin Invest. 2001; 108(9):1331-1339. (Biology). View Reference
  5. Liao F, Alderson R, Su J, Ullrich SJ, Kreider BL, Farber JM. STRL22 is a receptor for the CC chemokine MIP-3alpha. Biochem Biophys Res Commun. 1997; 236(1):212-217. (Biology). View Reference
  6. Liao F, Rabin RL, Smith CS, Sharma G, Nutman TB, Farber JM. CC-chemokine receptor 6 is expressed on diverse memory subsets of T cells and determines responsiveness to macrophage inflammatory protein 3 alpha. J Immunol. 1999; 162(1):186-194. (Biology). View Reference
  7. Lim HW, Lee J, Hillsamer P, Kim CH. Human Th17 cells share major trafficking receptors with both polarized effector T cells and FOXP3+ regulatory T cells. J Immunol. 2008; 180(1):122-129. (Biology). View Reference
  8. Power CA, Church DJ, Meyer A, et al. Cloning and characterization of a specific receptor for the novel CC chemokine MIP-3alpha from lung dendritic cells. J Exp Med. 1997; 186(6):825-835. (Biology). View Reference
  9. Yang D, Chertov O, Bykovskaia SN, et al. Beta-defensins: linking innate and adaptive immunity through dendritic and T cell CCR6. Science. 1999; 286(5439):525-528. (Biology). View Reference
  10. Zaballos A, Varona R, Gutierrez J, Lind P, Marquez G. Molecular cloning and RNA expression of two new human chemokine receptor-like genes. Biochem Biophys Res Commun. 1996; 227(3):846-853. (Biology). View Reference
View All (10) View Less
560620 Rev. 3

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