Anti gr1 clone rb6 8c5

Manufactured by BioXCell

Sourced in United States

Anti-Gr1 (clone RB6-8C5) is a monoclonal antibody that binds to the Gr-1 antigen, also known as Ly-6G and Ly-6C. The Gr-1 antigen is expressed on the surface of myeloid-derived suppressor cells (MDSCs), neutrophils, and some monocyte/macrophage subsets. This antibody can be used for the identification and isolation of these cell populations.

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Lab products found in correlation

Anti ly6g clone 1a8, by BioXCell (4 mentions) Clone ltf 2, by BioXCell (2 mentions) Ltf 2, by BioXCell (2 mentions) Rat igg, by Jackson ImmunoResearch (1 mentions) Anti pd 1 clone rmp1 14, by BioXCell (1 mentions) Anti ctla 4 clone 9h10, by BioXCell (1 mentions) Gemcitabine, by Selleck Chemicals (1 mentions) Anti cd8 clone 2, by BioXCell (1 mentions) Anti tim 3 clone rmt3 23, by BioXCell (1 mentions) Anti ox40 clone ox 86, by BioXCell (1 mentions) Anti lag 3 clone c9b7w, by BioXCell (1 mentions) Elispot, by R&D Systems (1 mentions) Anti cd8 clone 53, by BioXCell (1 mentions) Gp10025 33, by AnaSpec (1 mentions) Diphtheria toxin, by Merck Group (1 mentions) Poly 1 c, by GE Healthcare (1 mentions) Elisa max standard set, by BioLegend (1 mentions) Lps e coli o111 b4, by Merck Group (1 mentions) Amd3100, by Merck Group (1 mentions) Live dead fixable yellow dead cell stain kit, by Thermo Fisher Scientific (1 mentions)

18 protocols using anti gr1 clone rb6 8c5

Neutrophil and Alveolar Macrophage Depletion

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For neutrophil depletion, we followed an established protocol^{54 (link)}. Mice were injected i.p. with 125 μg of anti-GR1 (clone RB6-8C5, BioXCell, NH) (in 200 μl) per mouse 24 h before lung infection. Control mice received 125 μg of rat IgG (Jackson Immunoresearch). For depletion of alveolar macrophages, 100 μl CLL (purchased from http://clodronateliposomes.org/) was delivered intratracheally into mice 2 d before infection. Control mice received an equal volume of PBS-L.

Baral P., Umans B.D., Li L., Wallrapp A., Bist M., Kirschbaum T., Wei Y., Zhou Y., Kuchroo V.K., Burkett P.R., Yipp B.G., Liberles S.D, & Chiu I.M. (2018). Nociceptor sensory neurons suppress neutrophil and γδ T cell responses in bacterial lung infections and lethal pneumonia. Nature Medicine, 24(4), 417-426.

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Pharmacological Inhibition in Cancer Immunotherapy

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For in vivo pharmacological inhibition, gemcitabine (Selleck Chemicals) was dosed at 100mg/kg IP. SX-682 (Syntrix Pharmaceuticals) was dosed PO ad libitum (formulated concentration 714mg/kg feed); therapeutic plasma levels (range 0.5–10 μg/mL) were confirmed with this feed using LC/MS-MS. For ICT and Gr1/CD8-neutralizing antibody treatment, anti-PD1 (clone RMP1–14, BioXCell, BE0146), anti-CTLA4 (clone 9H10, BioXCell, BE0131), anti-TIM3 (clone RMT3–23, BioXCell, BE0115), anti-OX40 (clone OX-86, BioXCell, BE0031), anti-41BB (clone LOB12.3, BioXCell, BE0169), anti-LAG3 (clone C9B7W, BioXCell, BE0174), anti-CD8 (clone 2.43, BioXCell, BE0061) and anti-Gr1 (clone RB6–8C5, BioXCell, BE0075), antibodies (or their respective isotype IgG controls) were intraperitoneally administered at 200μg per injection three times per week. The duration of treatment was 4 weeks before endpoint analysis and survival analysis unless otherwise indicated.

Gulhati P., Schalck A., Jiang S., Shang X., Wu C.J., Hou P., Ruiz S.H., Soto L.S., Parra E., Ying H., Han J., Dey P., Li J., Deng P., Sei E., Maeda D.Y., Zebala J.A., Spring D.J., Kim M., Wang H., Maitra A., Moore D., Clise-Dwyer K., Wang Y.A., Navin N.E, & DePinho R.A. (2022). Targeting T cell checkpoints 41BB and LAG3 and myeloid cell CXCR1/2 results in anti-tumor immunity and durable response in pancreatic cancer. Nature cancer, 4(1), 62-80.

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Monocyte and Neutrophil Depletion Protocol

Cited 1 time

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To deplete monocytes and neutrophils, the mice were injected intraperitoneally with 500 μg of anti-Gr1 (clone RB6-8C5, BioXCell, West Lebanon, NH, USA) on days 0, 1, 2, 3 and 4 (ref. 13 (link)). To deplete neutrophils alone, the mice were injected intraperitoneally with 500 μg anti-Ly6G (clone 1A8, BioXCell) using the same schedule.

Singh T.P., Zhang H.H., Borek I., Wolf P., Hedrick M.N., Singh S.P., Kelsall B.L., Clausen B.E, & Farber J.M. (2016). Monocyte-derived inflammatory Langerhans cells and dermal dendritic cells mediate psoriasis-like inflammation. Nature Communications, 7, 13581.

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Antibody-mediated cell depletion

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Gr1⁺ or Ly6G⁺ cells were depleted by i.p. injection of 500 μg anti-Gr1 (clone RB6-8C5) or anti-Ly6G (clone 1A8) antibody (BioXCell, West Lebanon, NH), respectively. Control mice were injected with the equal amount of an isotype control antibody (clone MPC-11) (BioXCell). In experiments where tumor growth was studied in conjunction with cell depletion, tumor cells were inoculated 24 h after antibody injection. Six to seven mice were included in each group.

Deronic A., Leanderson T, & Ivars F. (2016). The anti-tumor effect of the quinoline-3-carboxamide tasquinimod: blockade of recruitment of CD11b+ Ly6Chi cells to tumor tissue reduces tumor growth. BMC Cancer, 16, 440.

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Deplete Immune Cells and Evaluate Adoptive T-cell Therapy

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To eliminate CD8⁺ T-cells or MDSC-like cells, LLC-bearing mice were treated with 400 μg anti-CD8 (clone 53.6.72, BioXcell) or 250 μg anti-Gr1 (clone RB6–8C5, BioXcell), respectively. Maintenance i.p. doses of the depleting antibodies were given every 3^rd day until tumor endpoint. In MDSC co-injection studies, 1×10⁶ tumor-MDSC, PMN-MDSC, or M-MDSC from Prkaa1^flox and Prkaa1^KO mice bearing LLC tumors were co-injected s.c. with 1×10⁶ LLC cells. For adoptive T-cell therapy (ACT), mice bearing B16 tumors for 6 days, received ACT with 1×10⁶ negatively sorted CD8⁺ pmel T-cells pre-activated for 48 hours with gp100_25–33 (AnaSpec). Ten days after pmel transfer, lymph nodes were activated for 24 hours with 1 μg/ml gp100_25–33 and tested for IFNγ expression using EliSpot (R&D systems).

Trillo-Tinoco J., Sierra R.A., Mohamed E., Cao Y., de Mingo-Pulido Á., Gilvary D.L., Anadon C.M., Costich T.L., Wei S., Flores E.R., Ruffell B., Conejo-Garcia J.R, & Rodriguez P.C. (2019). AMPK alpha-1 intrinsically regulates the function and differentiation of tumor myeloid-derived suppressor cells. Cancer research, 79(19), 5034-5047.

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Neutrophil Depletion in Scald Injury

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Mice were injected with 100 μg anti-Gr1 (clone RB6-8C5, BioXCell, West Lebanon, NH) in 100 μL sterile PBS i.p. on day 6 after scald injury. Spleens were harvested and neutrophil depletion confirmed by flow cytometry.

Beckmann N., Huber F., Hanschen M., St. Pierre Schneider B., Nomellini V, & Caldwell C.C. (2020). Scald Injury-Induced T Cell Dysfunction Can Be Mitigated by Gr1+ Cell Depletion and Blockage of CD47/CD172a Signaling. Frontiers in Immunology, 11, 876.

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Inflammatory Mediators and Cell Depletion

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For the induction of inflammation, lipopolysaccharides (LPS; E. coli, O111:B4) (Sigma), CpG-ODN (ODN1668; Hokkaido System Science), and Poly(I:C) (GE Healthcare Life Sciences) were used. For the depletion of monocytes and/or neutrophils, anti-Gr-1 (clone RB6-8C5, in-house purification) or anti-Ly6G (clone 1A8; BioXCell) was used. For the inhibition of MMP-9 activity and CXCR4, SB-3CT (Tokyo Chemical Industry) and AMD3100 (Sigma) were used respectively. Diphtheria toxin (DT) was purchased from Sigma. For the detection of IL-6 and TNF-alpha concentrations in serum, an ELISA MAX™ Standard Set was purchased from BioLegend. For analysis of cell surface marker expression, the following Abs were used: anti-CD11b-PE (clone M1/70), anti-CD62L-PE (clone MEL-14), anti-F4/80-PE (clone RM8), anti-C5aR-PE (clone 20/70), anti-MHC-II-PE (clone M5.114.15.2), anti-VCAM1-PE [clone 429 (MVCAM)], anti-Ly6G-PE (clone 1A8), anti-CXCR4-APC (L276F12), and anti-Treml4-PE (clone 16E5) were purchased from BioLegend. Anti-PD-L1-PE (clone MIH5) was purchased from Thermo Fisher Scientific. Anti-CD204-PE (clone REA148) was purchased from Miltenyi Biotec. Anti-CXCR2-APC (clone 242216) and anti-CCR2-APC (clone 475301R) were purchased from R&D Systems. Anti-CD131-PE (clone JORO50) was purchaced from BD Biosciences.

Shibuya T., Kamiyama A., Sawada H., Kikuchi K., Maruyama M., Sawado R., Ikeda N., Asano K., Kurotaki D., Tamura T., Yoneda A., Imada K., Satoh T., Akira S., Tanaka M, & Yotsumoto S. (2021). Immunoregulatory Monocyte Subset Promotes Metastasis Associated With Therapeutic Intervention for Primary Tumor. Frontiers in Immunology, 12, 663115.

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Antibody-mediated immune manipulation

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Anti-Ly6G (clone 1A8, #BP0075-1), anti-Gr1 (clone RB6-8C5, #BE0075), anti-rat Kappa immunoglobulin (clone MAR 18.5, #BE0122) and corresponding isotype controls (#BP0290 and #BP0089) were all purchased from Bio X Cell and injected intraperitoneally following the schemes and dosage that are presented in the Figures or legends. When mice were sequentially injected with two antibodies, we respected 2 h delay between injections.

Boivin G., Faget J., Ancey P.B., Gkasti A., Mussard J., Engblom C., Pfirschke C., Contat C., Pascual J., Vazquez J., Bendriss-Vermare N., Caux C., Vozenin M.C., Pittet M.J., Gunzer M, & Meylan E. (2020). Durable and controlled depletion of neutrophils in mice. Nature Communications, 11, 2762.

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MDSC Depletion in Tumor-Bearing Mice

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When tumors were palpable, mice were injected (i.p.) with anti-Gr1 (clone RB6-8C5, BioXCell) or control antibody (clone LTF-2, BioXCell) (200µg/injection, in 100µl PBS) every 48 hours. MDSC depletion was confirmed in the peripheral blood of recipient mice. Blood sample was recovered in heparin containing tubes. After red blood cell lysis by ACK, nucleated cells were incubated for 20 minutes at 4°C with blocking antibody (24G2, 1mg/ml, 1/100). Cells were stained for 30 minutes at 4°C with the following antibodies: PECy7 anti-CD45 (Clone 30-F11, BD Pharmingen, 1/800), eF450 anti-CD11b (Clone M1/70, eBioscience, 1/100), PerCPCy5.5 anti-Gr1 (clone RB6-8C5, BD Pharmingen, 1/100), PE anti-CD11c (Clone N418, eBioscience, 1/100) and LIVE/DEAD® Fixable Yellow Dead Cell Stain Kit (Life Technologies, L34959, 1/100). Staining was assessed with a FACS Fortessa cytometer, and flow cytometry data were analyzed using FACSDiva (BD Biosciences) or FlowJO (Tree Star) software.

Dajon M., Iribarren K., Petitprez F., Marmier S., Lupo A., Gillard M., Ouakrim H., Victor N., Vincenzo D.B., Joubert P.E., Kepp O., Kroemer G., Alifano M., Damotte D, & Cremer I. (2018). Toll like receptor 7 expressed by malignant cells promotes tumor progression and metastasis through the recruitment of myeloid derived suppressor cells. Oncoimmunology, 8(1), e1505174.

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Modulating MDSC with Peptibody Treatment

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Anti-Gr-1 (clone RB6-8C5) and isotype control (clone LTF-2) were purchased from BioXcell and dosed at 200 μg/mouse (i.p.) every other day. 15 μg of endotoxin-free plasmids for irrelevant control peptibody (Irr-pep) and MDSC-specific Pep-H6 peptibody were injected into mice through tail vein using the established protocol (21 (link)) in TransIT-EE Delivery Solution (Mirus Bio LLC) every 4 days. SB225002 (Cayman Chemical) in DMSO was diluted in vehicle (0.9%NaCl, 0.3%Tween 80) for in vivo administration every other day (5 mg/kg).

Wang G., Lu X., Dey P., Deng P., Wu C.C., Jiang S., Fang Z., Zhao K., Konaparthi R., Hua S., Zhang J., Li-Ning-Tapia E.M., Kapoor A., Wu C.J., Patel N.B., Guo Z., Ramamoorthy V., Tieu T.N., Heffernan T., Zhao D., Shang X., Khadka S., Hou P., Hu B., Jin E.J., Yao W., Pan X., Ding Z., Shi Y., Li L., Chang Q., Troncoso P., Logothetis C.J., McArthur M.J., Chin L., Wang Y.A, & DePinho R.A. (2015). Targeting YAP-dependent MDSC infiltration impairs tumor progression. Cancer discovery, 6(1), 80-95.

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