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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.
BD Pharmingen™ PE-Cy™7 Rat Anti-Mouse Ly-6G and Ly-6C
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- Since applications vary, each investigator should titrate the reagent to obtain optimal results.
- An isotype control should be used at the same concentration as the antibody of interest.
- 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.
- 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.
- For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
- 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).
- 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.
- Cy is a trademark of GE Healthcare.
- Please refer to http://regdocs.bd.com to access safety data sheets (SDS).
- Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
配套商品
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.
研发参考 (13)
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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). 查看参考
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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). 查看参考
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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). 查看参考
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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). 查看参考
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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). 查看参考
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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). 查看参考
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Lagasse E, Weissman IL. Flow cytometric identification of murine neutrophils and monocytes. J Immunol Methods. 1996; 197(1-2):139-150. (Biology). 查看参考
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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). 查看参考
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Roederer M, Kantor AB, Parks DR, Herzenberg LA. Cy7PE and Cy7APC: bright new probes for immunofluorescence. Cytometry. 1996; 24(3):191-197. (Methodology). 查看参考
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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). 查看参考
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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). 查看参考
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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). 查看参考
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