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Ly6g apc cy7

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Ly6G-APC-Cy7 is a fluorochrome-conjugated antibody that binds to the Ly6G antigen. It is commonly used in flow cytometry applications to identify and analyze granulocytes, particularly neutrophils, within a sample.

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Lab products found in correlation

Fc block, by BD (2 mentions) Rbc lysis buffer, by Thermo Fisher Scientific (2 mentions) Collagenase, by Merck Group (2 mentions) F4 80 apc facs antibody, by Bio-Rad (2 mentions) Cd11b pe cy7, by BD (2 mentions) Ly6c fitc, by BD (2 mentions) Cd45 percp, by BD (2 mentions) Cd11b apc, by BD (1 mentions) Fitc annexin 5 apoptosis detection kit, by BD (1 mentions) Siglec f, by BD (1 mentions) Facsaria cell sorter, by BD (1 mentions) Facsdiva software version 6, by BD (1 mentions) Bx60 fluorescence microscope, by Olympus (1 mentions) Shandon cytospin 3 cytocentrifuge, by Thermo Fisher Scientific (1 mentions) Cd11b pe cf594, by BD (1 mentions) Rpmi 1640, by Thermo Fisher Scientific (1 mentions) Il 2 pe, by BD (1 mentions) Cd80 pe, by BD (1 mentions) Cd86 apc, by BD (1 mentions) Ifn γ apc, by BD (1 mentions)

Close spelling variants of this product

APC-Cy7 (76 citations) Ly6G-APC (23 citations) Anti-Ly6G-APC-Cy7 (4 citations) Anti-Ly6G-APC-Cy7 (1A8, (3 citations) Ly6G-APC Cy7 (1A8), (2 citations)

13 protocols using ly6g apc cy7

1

Differential Leukocyte Apoptosis Analysis in BALF

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After being filtered through a 100μm nylon mesh, BALF was centrifuged at 400 x g at 4°C for 7 minutes and then discarded the supernatant. The cells were incubated with 200 μL ACK lysis at room temperature for 2 min and then 1 mL cold DPBS was to stop the reaction. After being centrifuged at 400 x g at 4°C for 7 minutes, the supernatant was discarded and the cell pellet was resuspended with binding buffer (5% FBS) at 2 x 10^6/mL. Incubate the cells with the fluorescent dye-conjugated mouse antibodies at 4°C for 30 min and avoid direct light. Neutrophils in BALF were marked as LY-6G + (APC-CY7; BD Biosciences, NJ, USA) and CD 11b + (APC; BD Biosciences, NJ, USA); eosinophils in BALF were marked as LY-6G – (APC-CY7; BD Biosciences, NJ, USA) and Siglec-F + (BV421; BD Biosciences, NJ, USA). After being washed with 1mL FBS, the cells were centrifuged at 400 x g at 4°C for 7 minutes before the detection of apoptosis. The FITC Annexin V Apoptosis Detection Kit was purchased from BD Pharmingen (NJ, USA), and the procedures followed the manufacturer’s protocol. Cells were analyzed via a BD Biosciences flow cytometer, and the results were analyzed using FlowJo software (Version 10.8.1).
Xiao S., Zhou Y., Wang Q, & Yang D. (2022). Ketamine Attenuates Airway Inflammation via Inducing Inflammatory Cells Apoptosis and Activating Nrf2 Pathway in a Mixed-Granulocytic Murine Asthma Model. Drug Design, Development and Therapy, 16, 4411-4428.
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2

Isolation and FACS Analysis of Stromal Vascular Cells

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Stromal vascular cells (SVCs) isolation and FACS analyses were performed as previously described61 ,62 . Briefly, epididymal fat pads were weighed, rinsed three times in PBS, and then minced in 1% BSA PBS. Tissue suspensions were digested with collagenase (1 mg ml−1, Sigma-Aldrich) for 30 minutes. After centrifugation at 500 g for 5 min, the pellet containing SVCs was incubated with red blood cell (RBC) lysis buffer (eBioscience) for 5 min followed by another centrifugation (300 g, 5 min) and resuspension in 1% BSA PBS. SVCs were incubated with Fc Block (BD Biosciences) for 20 min at 4 °C before staining with fluorescently labeled primary antibodies or control IgGs for 30 min at 4 °C. F4/80-APC FACS antibody was purchased from AbD Serotec (Raleigh, NC); FITC-Cd11b, PE-Cd11c and APC- Cy7-Ly6G antibodies were from BD Biosciences.
Li P., Oh D.Y., Bandyopadhyay G., Lagakos W.S., Talukdar S., Osborn O., Johnson A., Chung H., Maris M., Ofrecio J.M., Taguchi S., Lu M, & Olefsky J.M. (2015). LTB4 causes macrophage–mediated inflammation and directly induces insulin resistance in obesity. Nature medicine, 21(3), 239-247.
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3

Isolation and FACS Analysis of Stromal Vascular Cells

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Stromal vascular cells (SVCs) isolation and FACS analyses were performed as previously described61 ,62 . Briefly, epididymal fat pads were weighed, rinsed three times in PBS, and then minced in 1% BSA PBS. Tissue suspensions were digested with collagenase (1 mg ml−1, Sigma-Aldrich) for 30 minutes. After centrifugation at 500 g for 5 min, the pellet containing SVCs was incubated with red blood cell (RBC) lysis buffer (eBioscience) for 5 min followed by another centrifugation (300 g, 5 min) and resuspension in 1% BSA PBS. SVCs were incubated with Fc Block (BD Biosciences) for 20 min at 4 °C before staining with fluorescently labeled primary antibodies or control IgGs for 30 min at 4 °C. F4/80-APC FACS antibody was purchased from AbD Serotec (Raleigh, NC); FITC-Cd11b, PE-Cd11c and APC- Cy7-Ly6G antibodies were from BD Biosciences.
Li P., Oh D.Y., Bandyopadhyay G., Lagakos W.S., Talukdar S., Osborn O., Johnson A., Chung H., Maris M., Ofrecio J.M., Taguchi S., Lu M, & Olefsky J.M. (2015). LTB4 causes macrophage–mediated inflammation and directly induces insulin resistance in obesity. Nature medicine, 21(3), 239-247.
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4

Immunophenotyping of Myeloid Cells in Dams

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Dams at 10.5 dpc and 18.5 dpc were euthanized and peripheral blood was obtained by cardiac puncture (n = 10 each). Peripheral leukocytes were incubated with the CD16/CD32 monoclonal antibody (mAb) (FcγIII/II receptor; BD Biosciences, San Jose, CA) followed by staining using CD11b-PECF594, Ly6G-APC-Cy7 and F4/80-PE mAbs (BD Biosciences), after which the cell suspensions underwent intracellular staining with either OVA-FITC antibody or rabbit IgG-FITC isotype control. Cells were resuspended in 500 μL of FACS buffer and sorted using a BD FACSAria cell sorter (BD Biosciences) and BD FACSDiva Software Version 6.1.3. The sorted CD11b+Ly6G+OVA+ or CD11b+F4/80+OVA+ cells were then resuspended with 200 μL of FACS buffer. Cytospin slides of sorted cells were prepared using Fisherbrand Superfrost microscope slides (Thermo Fisher Scientific) and a Shandon Cytospin 3 cytocentrifuge (Thermo Fisher Scientific) at 800 rpm for 5 min. After centrifugation, all slides were washed with 1X PBS and the cells were fixed with 4% paraformaldehyde for 20 min. After fixation, the slides were washed with 1X PBS, dried, and mounted using ProLong Diamond Antifade Mountant with DAPI. Images were obtained with an Olympus BX60 fluorescence microscope at 40x magnification with digital zoom.
Arenas-Hernandez M., Romero R., Gershater M., Tao L., Xu Y., Garcia-Flores V., Pusod E., Miller D., Galaz J., Motomura K., Schwenkel G., Para R, & Gomez-Lopez N. (2021). Specific Innate Immune Cells Uptake Fetal Antigen and Display Homeostatic Phenotypes in the Maternal Circulation. Journal of leukocyte biology, 111(3), 519-538.
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5

Multiparameter Flow Cytometry Analysis

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RPMI 1640, FBS, penicillin, and streptomycin were obtained from Invitrogen (Grand Island, NY). The liver dissociation kit, recombinant murine was from Miltenyi Biotec. Phorbol 12-myristate 13-acetate (PMA), brefeldin A, ionomycin, CD3, CD28, and dimethyl sulfoxide (DMSO) were purchased from Sigma-Aldrich (St. Louis, MO). We obtained the following fluorescein-conjugated anti-mouse antibodies from eBioscience (San Diego, CA), Biolegend (San Diego, CA) and BD: CD3e-APC-Cy7 (145-2C11), CD4-PerCP-Cy5.5 (RM4-5), CD8a-PE (53-6.7), ICOS-PE-Cy7 (C398.4A), CD69-BV421 (MIH5), CD19- PE-Cy5(6D5), CD138-PE-Cy7 (281-2), B220-APC-Cy7 (RA3632), CD80-PE (16-10A1), CD86-APC (GL1), CD3e-FITC (145-2C1), CD11b- PE-Cy7 (M1/70), Ly-6C-PerCP-Cy5.5 (HK1.4), F4/80- APC-Cy7 (3M8), CD192/CCR2-BV421 (SA203G11), CX3CR1-APC (SA011F11), CD135-BV421 (A2F10.1), CD11c- PerCP-Cy5.5 (HL3), Gr-1-FITC (RB6-8C5), Ly-6G- APC-Cy7 (1A8), CD287/TLR7-PE (A94B10), CD103-PE (M290), IFN-γ-APC (XMG1.2), IL-4-PE (11B11), IL-2-PE (JES6-5H4), IL-6-APC (MP5-20F3), IL-10-PE (JES5-16E3), IL-13-eFlour450 (ebio13A), IL-17-PE (TC11-18H10.1)), IL-21-APC (FFA21) and their corresponding isotype controls.
Feng Y., Xie H., Shi F., Chen D., Xie A., Li J., Fang C., Wei H., Huang H., Pan X., Tang X, & Huang J. (2021). Roles of TLR7 in Schistosoma japonicum Infection-Induced Hepatic Pathological Changes in C57BL/6 Mice. Frontiers in Cellular and Infection Microbiology, 11, 754299.
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6

Comprehensive Immune Cell Profiling

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CD11b-PE-Cy7 (Thermo Fisher Scientific, 25-0112-82), Rat IgGk Isotype Control-PE-Cy7 (Thermo Fisher Scientific, 25-4031-82) CD45-e450 (Thermo Fisher Scientific, 48-0451-82), Rat IgG2b Isotype Control-e450 (Thermo Fisher Scientific, 48-4031-82), CD45.1-e450 (Biolegend, 110721), Mouse IgG2ak Isotype Control-e450 (Biolegend, 400235), CD45.2-APC (Biolegend, 109813), Mouse IgG2ak Isotype Control-APC (Biolegend, 400221), Ly6G-APC-Cy7 (BD Biosciences, 560600), Rat IgG2a Isotype Control-APC-Cy7 (BD Biosciences, 552770), CD11c-PerCP-Cy5.5 (Thermo Fisher Scientific, 45-0114-82), Arm Ham IgG Isotype Control-PerCP-Cy5.5 (Thermo Fisher Scientific, 45-4888-80), Ly6C-FITC (BD Biosciences, 561085), Rat IgM Isotype Control-FITC (BD Biosciences, 553942). All antibodies were used at a working concentration of 1:100 except for CD11b (1:200).
Kalinski A.L., Yoon C., Huffman L.D., Duncker P.C., Kohen R., Passino R., Hafner H., Johnson C., Kawaguchi R., Carbajal K.S., Jara J.S., Hollis E., Geschwind D.H., Segal B.M, & Giger R.J. (2020). Analysis of the immune response to sciatic nerve injury identifies efferocytosis as a key mechanism of nerve debridement. eLife, 9, e60223.
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7

Comprehensive Immune Profiling of CT26 Tumors

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CT26 tumors were digested using a gentleMACS dissociator and a murine tumor dissociation kit (Miltenyi Biotec). Absolute viable cell counts were determined by propidium iodide staining and analysed on the MACSQuant analyzer. Cells from CT26 tumors and splenocytes were stained with fixable LIVE/DEAD blue (Life Technologies) and incubated with anti-mouse CD16/CD32 (eBioscience) prior to addition of anti-mouse: CD8-Pe-Cy7 (clone 53–6.7); CD3-eFluor 450 (clone 17A2); CD11c-PE (clone N418); CD86-FITC (clone GL1); PDCA1-APC (clone 129c) (eBioscience); PD-L1-BV421 (clone 10F.9G2); I-A/I-E (MHCII) (clone M5/114.15.2); B220-BV605 (clone RA3-6B2) (Biolegend); CD45-BV785 (clone 30F11); CD4-BUV395 (clone GK1.5); CD11b-BUV395 (clone M1/70); Ly6G-APC-Cy7 (clone 1A8); Ly6C-PerCP-Cy5.5 (clone AL-21) (BD Biosciences). For intracellular staining, cells were permeabilized using Foxp3 / Transcription Factor Staining Buffer Set (eBioscience) and incubated with anti-mouse Foxp3-PE (clone FJK-16S) and Ki67-eFluor 660 (clone SolA15) (eBioscience). Stained cells were fixed in 3.7% formaldehyde and analyzed using a BD LSRFortessa (BD Bioscience). Data analysis was performed using FlowJo (FlowJo LLC). 8 mice per treatment group were included in all flow cytometry analyses.
Poon E., Mullins S., Watkins A., Williams G.S., Koopmann J.O., Di Genova G., Cumberbatch M., Veldman-Jones M., Grosskurth S.E., Sah V., Schuller A., Reimer C., Dovedi S.J., Smith P.D., Stewart R, & Wilkinson R.W. (2017). The MEK inhibitor selumetinib complements CTLA-4 blockade by reprogramming the tumor immune microenvironment. Journal for Immunotherapy of Cancer, 5, 63.
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8

Isolation and Analysis of Spinal Cord Cells

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CXCR1CreER; Rosa26tdTom mice were injected with a terminal dose of ketamine and xylazine. Mice were flushed with PBS, their spinal cords were removed, and the LPC lesion sites were microdissected 7 days after LPC. Tissue was diced and digested in a solution containing papain (1.54 μg/ml; Worthington, Lakewood, NJ), l-cysteine (360 μg/ml; Sigma-Aldrich) and DNAseI (703 μg/ml; Roche) for 25 min at 37°C. Tissue was then dissociated by trituration in 20% fetal bovine serum (FBS) and filtered through a 70-μm filter before staining. For blood flow cytometry, blood was collected through cardiac puncture and placed in a tube with heparin before diluting sample with Hank’s buffered salt solution (HBSS). Cells were exposed to Fc block (1:100, αCD16/32; BD Pharmingen) for 30 min at 4°C, and the following antibodies were used to stain tissue at 1:50 for 45 min at 4°C: CD45-PerCP (BD Pharmingen; clone 30-F11), CD11b-FITC (BD Pharmingen; clone M1/70), Ly6C-V450 (BD Horizon; clone Al-21), and Ly6G-APC-Cy7 (BD Pharmingen; clone 1A8). For blood staining, red blood cells were subsequently lysed by addition of BD FACS Lysing Solution (BD Biosciences) while rocking for 12 min at room temperature. Cells were washed and fixed in 1% buffered formalin for flow cytometry analysis (conducted by University of Calgary Flow Cytometry Core Facility). Data were analyzed using FlowJo 10.0.8 (FlowJo).
Plemel J.R., Stratton J.A., Michaels N.J., Rawji K.S., Zhang E., Sinha S., Baaklini C.S., Dong Y., Ho M., Thorburn K., Friedman T.N., Jawad S., Silva C., Caprariello A.V., Hoghooghi V., Yue J., Jaffer A., Lee K., Kerr B.J., Midha R., Stys P.K., Biernaskie J, & Yong V.W. (2020). Microglia response following acute demyelination is heterogeneous and limits infiltrating macrophage dispersion. Science Advances, 6(3), eaay6324.
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9

Isolation and Characterization of Mouse Cardiac Immune Cells

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The mouse heart was perfused, excised, minced, digested with type I collagenase, mechanically disrupted, and filtered through a 70 μm cell strainer to get a single-cell suspension (21 (link)). Cells were collected by centrifugation, subjected to live-dead dye (LIVE/DEAD Fixable Yellow Dead Cell Stain Kit, Invitrogen, L34968) and surface antibody staining (fluorescent conjugated primary antibodies; BioLegend), and analyzed by flow cytometry. Cells were also sorted by flow cytometry to collect live CD45+ cells for transcriptomic studies. The following fluorescent antibodies were used in this study: CD45-PerCP (BD Biosciences, 561047), CD11b-AF488 (BD Biosciences, 557672), Ly6G-APC-Cy7 (BD Biosciences, 560600), F4/80-BV421 (BioLegend, 123137), and CD45-FITC (BioLegend, 103108).
Liao X., Chang E., Tang X., Watanabe I., Zhang R., Jeong H.W., Adams R.H, & Jain M.K. (2022). Cardiac macrophages regulate isoproterenol-induced Takotsubo-like cardiomyopathy. JCI Insight, 7(3), e156236.
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10

Tumor-Associated Macrophages Profiling

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Tumors were harvested and processed to obtain single cell suspensions for determination of TAM burden by flow cytometry. Briefly, minced tumor tissues were digested with collagenase IV, dispase II, and DNAse I (Sigma). Single cell suspension was then treated with ACK lysing buffer (Lonza) to remove red blood cells, and immune cells were isolated using a Percoll (GE Healthcare) gradient [28 (link)]. Cells were stained with CD45 PE, CD11b PerCP, F4/80 Pacific Blue, Ly6C FITC, Ly6G APC-Cy7 (BD Biosciences), and the Aqua Live/Dead viability dye (Molecular Probes). Cells were then analyzed on the LSRII four-laser flow cytometer (BD Biosciences) using BD FACSDiva software (version 6.0) and FlowJo (version 7.2.5; BD Biosciences). TAMs were defined as percentage of CD45 cells that were CD11b+, Ly6G−, Ly6C−, and F4/80+ [29 (link)].
Starosolski Z., Courtney A.N., Srivastava M., Guo L., Stupin I., Metelitsa L.S., Annapragada A, & Ghaghada K.B. (2021). A Nanoradiomics Approach for Differentiation of Tumors Based on Tumor-Associated Macrophage Burden. Contrast Media & Molecular Imaging, 2021, 6641384.
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