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PE-Cy™7 Rat Anti-Mouse Ly-6G and Ly-6C
PE-Cy™7 Rat Anti-Mouse Ly-6G and Ly-6C

Two-parameter flow cytometric analysis of Ly-6G and/or Ly-6C expression on bone-marrow myeloid cells. BALB/c bone-marrow leucocytes were stained with either PE-Cy™7 Rat IgG2b, κ Isotype Control (Cat. No. 552849; Left Plot) or PE-Cy™7 Rat Anti-Mouse Ly-6G and Ly-6C antibody (Cat. No. 552985/565033; Right Plot). Two-parameter flow cytometric contour plots showing the correlated expression of Ly-6G and Ly-6C (or Ig Isotype control staining) versus side light-scatter (SSC) signals were derived from gated events with the forward and side light-scatter characteristics of bone-marrow leucocyte populations. Please note that the population of cells having the lowest side scattered-light (SSC) characteristics (erythroid and lymphoid cells) shows little expression of Ly-6G/Ly-6C, while most of the leucocytes with moderate-to-high SSC (myeloid cells) are Ly-6G/Ly-6C-positive. Flow cytometric analysis was performed using a BD FACSCalibur™ Flow Cytometry System.

Two-parameter flow cytometric analysis of Ly-6G and/or Ly-6C expression on bone-marrow myeloid cells. BALB/c bone-marrow leucocytes were stained with either PE-Cy™7 Rat IgG2b, κ Isotype Control (Cat. No. 552849; Left Plot) or PE-Cy™7 Rat Anti-Mouse Ly-6G and Ly-6C antibody (Cat. No. 552985/565033; Right Plot). Two-parameter flow cytometric contour plots showing the correlated expression of Ly-6G and Ly-6C (or Ig Isotype control staining) versus side light-scatter (SSC) signals were derived from gated events with the forward and side light-scatter characteristics of bone-marrow leucocyte populations. Please note that the population of cells having the lowest side scattered-light (SSC) characteristics (erythroid and lymphoid cells) shows little expression of Ly-6G/Ly-6C, while most of the leucocytes with moderate-to-high SSC (myeloid cells) are Ly-6G/Ly-6C-positive. Flow cytometric analysis was performed using a BD FACSCalibur™ Flow Cytometry System.

Product Details
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BD Pharmingen™
Ly6c, Lymphocyte antigen 6C2; Lymphocyte antigen 6G, Ly6g, Gr-1
Mouse (QC Testing)
Rat IgG2b, κ
Not Reported
Flow cytometry (Routinely Tested)
0.2 mg/ml
AB_2739049
Aqueous buffered solution containing protein stabilizer 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.

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. 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. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
  6. 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).
  7. 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.
  8. Cy is a trademark of GE Healthcare.
  9. Please refer to http://regdocs.bd.com to access safety data sheets (SDS).
  10. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
565033 Rev. 2
Antibody Details
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RB6-8C5

The RB6-8C5 monoclonal antibody recognizes a common epitope on Ly-6G and Ly-6C, previously known as the myeloid differentiation antigen Gr-1. In the bone marrow, the level of antigen expression is directly correlated with granulocyte differentiation and maturation. The antigen is also expressed on the monocyte lineage in the bone marrow, but not on erythroid cells. In the periphery, RB6-8C5 antibody recognizes granulocytes (neutrophils and eosinophils) and monocytes. The RB6-8C5 antibody is a component of the "lineage cocktail" used in studies of hematopoietic cell lineages. The 1A8 antibody (Cat. No. 551461) specifically recognizes Ly-6G, but not Ly-6C.

Based on comparison of the staining patterns given by 1A8 versus RB6-8C5 antibodies on total blood leucocytes, it is evident that the 1A8 antibody stains the RB6-8C5-bright population, corresponding to Ly-6G-expressing granulocytes; whereas, the RB6-8C5-dim population is 1A8-negative and corresponds to Ly-6C-expressing lymphocytes and monocytes. Please refer to the Technical Data Sheets for Cat. No. 551459 and 553128 for more detailed information.

        

565033 Rev. 2
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
565033 Rev.2
Citations & References
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Development References (13)

  1. Brummer E, Sugar AM, Stevens DA. Immunological activation of polymorphonuclear neutrophils for fungal killing: studies with murine cells and blastomyces dermatitidis in vitro. J Leukoc Biol. 1984; 36(4):505-520. (Clone-specific: Cytotoxicity). View Reference
  2. Conlan JW, North RJ. Neutrophils are essential for early anti-Listeria defense in the liver, but not in the spleen or peritoneal cavity, as revealed by a granulocyte-depleting monoclonal antibody. J Exp Med. 1994; 179(1):259-268. (Clone-specific: Depletion). View Reference
  3. Czuprynski CJ, Brown JF, Maroushek N, Wagner RD, Steinberg H. Administration of anti-granulocyte mAb RB6-8C5 impairs the resistance of mice to Listeria monocytogenes infection. J Immunol. 1994; 152(4):1836-1846. (Clone-specific: Depletion, Western blot). View Reference
  4. Fleming TJ, Fleming ML, Malek TR. Selective expression of Ly-6G on myeloid lineage cells in mouse bone marrow. RB6-8C5 mAb to granulocyte-differentiation antigen (Gr-1) detects members of the Ly-6 family. J Immunol. 1993; 151(5):2399-2408. (Clone-specific: Immunoprecipitation, Inhibition). View Reference
  5. Gumley TP, McKenzie IF, Sandrin MS. Tissue expression, structure and function of the murine Ly-6 family of molecules. Immunol Cell Biol. 1995; 73(4):277-296. (Biology). View Reference
  6. Hestdal K, Ruscetti FW, Ihle JN, et al. Characterization and regulation of RB6-8C5 antigen expression on murine bone marrow cells. J Immunol. 1991; 147(1):22-28. (Biology). View Reference
  7. Jutila MA, Kroese FG, Jutila KL, et al. Ly-6C is a monocyte/macrophage and endothelial cell differentiation antigen regulated by interferon-gamma. Eur J Immunol. 1988; 18(11):1819-1826. (Clone-specific: Western blot). View Reference
  8. Lagasse E, Weissman IL. Flow cytometric identification of murine neutrophils and monocytes. J Immunol Methods. 1996; 197(1-2):139-150. (Biology). View Reference
  9. Lewinsohn DM, Bargatze RF, Butcher EC. Leukocyte-endothelial cell recognition: evidence of a common molecular mechanism shared by neutrophils, lymphocytes, and other leukocytes. J Immunol. 1987; 138(12):4313-4321. (Biology). View Reference
  10. Roederer M, Kantor AB, Parks DR, Herzenberg LA. Cy7PE and Cy7APC: bright new probes for immunofluorescence. Cytometry. 1996; 24(3):191-197. (Methodology). View Reference
  11. Stoppacciaro A, Melani C, Parenza M, et al. Regression of an established tumor genetically modified to release granulocyte colony-stimulating factor requires granulocyte-T cell cooperation and T cell-produced interferon gamma. J Exp Med. 1993; 178(1):151-161. (Clone-specific: Depletion, Immunohistochemistry). View Reference
  12. Tepper RI, Coffman RL, Leder P. An eosinophil-dependent mechanism for the antitumor effect of interleukin-4. Science. 1992; 257(5069):548-551. (Clone-specific: Depletion). View Reference
  13. Tumpey TM, Chen SH, Oakes JE, Lausch RN. Neutrophil-mediated suppression of virus replication after herpes simplex virus type 1 infection of the murine cornea. J Virol. 1996; 70(2):898-904. (Clone-specific: Depletion). View Reference
View All (13) View Less
565033 Rev. 2

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