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PE Rat Anti-Mouse Ly-6G and Ly-6C
PE Rat Anti-Mouse Ly-6G and Ly-6C

Representative staining of peripheral blood leukocytes with PE-conjugated antibody RB6-8C5. C57BL/6 whole blood was stained with PE-conjugated RB6-8C5 (anti-Ly-6G and Ly-6C) and FITC-conjugated 1A8 (anti-mouse Ly-6G, Cat. No. 551460) monoclonal antibodies in the presence of Mouse BD Fc Block™ purified anti-mouse CD16/CD32 mAb 2.4G2 (Cat. No. 553141/553142, left panel). Erythrocytes were lysed (BD Pharm Lyse™ lysis buffer, Cat. No. 555899) and non-viable leukocytes were excluded by staining with propidium iodide. The left panel demonstrates that mAb 1A8 stains the RB6-8C5hi population, corresponding to Ly-6G-expressing granulocytes; whereas,  the RB6-8C5lo population is 1A8-negative and corresponds to Ly-6C-expressing lymphocytes and monocytes. Backgating of the RB6-8C5+/1A8- population (R1) onto the light-scatter profile (right panel) indicates that this population falls within the monocyte region of the light-scatter profile. The RB6-8C5+/1A8+ population (R2) falls within the granulocyte region of the light-scatter profile (right panel). Flow cytometry was performed on a BD FACSCalibur™ flow cytometry system.

Representative staining of peripheral blood leukocytes with PE-conjugated antibody RB6-8C5. C57BL/6 whole blood was stained with PE-conjugated RB6-8C5 (anti-Ly-6G and Ly-6C) and FITC-conjugated 1A8 (anti-mouse Ly-6G, Cat. No. 551460) monoclonal antibodies in the presence of Mouse BD Fc Block™ purified anti-mouse CD16/CD32 mAb 2.4G2 (Cat. No. 553141/553142, left panel). Erythrocytes were lysed (BD Pharm Lyse™ lysis buffer, Cat. No. 555899) and non-viable leukocytes were excluded by staining with propidium iodide. The left panel demonstrates that mAb 1A8 stains the RB6-8C5hi population, corresponding to Ly-6G-expressing granulocytes; whereas,  the RB6-8C5lo population is 1A8-negative and corresponds to Ly-6C-expressing lymphocytes and monocytes. Backgating of the RB6-8C5+/1A8- population (R1) onto the light-scatter profile (right panel) indicates that this population falls within the monocyte region of the light-scatter profile. The RB6-8C5+/1A8+ population (R2) falls within the granulocyte region of the light-scatter profile (right panel). Flow cytometry was performed on 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_394644
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 R-PE under optimum conditions, and unconjugated antibody and free PE were removed.

Recommended Assay Procedures

Mouse BD Fc Block™ purified anti-mouse CD16/CD32 mAb 2.4G2 (Cat. No. 553141/553142) may help to reduce non-specific binding to cells bearing Fcγ-receptors.

Product Notices

  1. Since applications vary, each investigator should titrate the reagent to obtain optimal results.
  2. 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.
  3. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
  4. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
553128 Rev. 21
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.

553128 Rev. 21
Format Details
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PE
R-Phycoerythrin (PE), is part of the BD family of Phycobiliprotein dyes. This fluorochrome is a multimeric fluorescent phycobiliprotein with excitation maximum (Ex Max) of 496 nm and 566 nm and an emission maximum (Em Max) at 576 nm. PE is designed to be excited by the Blue (488 nm), Green (532 nm) and Yellow-Green (561 nm) lasers and detected using an optical filter centered near 575 nm (e.g., a 575/26-nm bandpass filter). As PE is excited by multiple lasers, this can result in cross-laser excitation and fluorescence spillover on instruments with various combinations of Blue, Green, and Yellow-Green lasers. Please ensure that your instrument’s configurations (lasers and optical filters) are appropriate for this dye.
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PE
Yellow-Green 488 nm, 532 nm, 561 nm
496 nm, 566 nm
576 nm
553128 Rev.21
Citations & References
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Development References (8)

  1. 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
  2. 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
  3. 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
  4. 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
  5. Lagasse E, Weissman IL. Flow cytometric identification of murine neutrophils and monocytes. J Immunol Methods. 1996; 197(1-2):139-150. (Biology). View Reference
  6. Osawa M, Tokumoto Y, Nakauchi H. Hematopoietic stem cells. In: Herzenberg LA, Weir DM, Blackwell C, ed. Weir's Handbook of Experimental Immunology, 5th Edition. Cambridge: Blackwell Science; 1996:66.1-66.5.
  7. 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
  8. 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
View All (8) View Less
553128 Rev. 21

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