Cd45 apc cy7

Manufactured by BioLegend

Sourced in United States, Germany

CD45-APC-Cy7 is a fluorescently-conjugated antibody that binds to the CD45 protein, which is expressed on the surface of various hematopoietic cells. It can be used for the identification and analysis of these cell populations in flow cytometry applications.

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Lab products found in correlation

Cd11b pe cy7, by BioLegend (12 mentions) Lsrii flow cytometer, by BD (8 mentions) Trustain fcx, by BioLegend (7 mentions) Cytoflex, by Beckman Coulter (6 mentions) Cd3 pe cy7, by BioLegend (6 mentions) Ly6g bv510, by BioLegend (5 mentions) Facscanto 2, by BD (5 mentions) Cd4 apc, by BioLegend (5 mentions) Cd206 fitc, by BioLegend (5 mentions) Ly6g fitc, by BioLegend (5 mentions) Cd11c pe cy7, by BioLegend (5 mentions) Cd11b pe cy7, by Thermo Fisher Scientific (5 mentions) Facscalibur, by BD (5 mentions) F4 80 pe, by BioLegend (4 mentions) Cd4 percp cy5, by BioLegend (4 mentions) Ly6g percp cy5, by BioLegend (4 mentions) Fc block, by BD (4 mentions) Cd3 fitc, by BioLegend (4 mentions) Fetal bovine serum (fbs), by Thermo Fisher Scientific (4 mentions) Cd16 32, by BioLegend (4 mentions)

Spelling variants of this product

APC-Cy7 (106 citations) CD45-APC (97 citations) Anti-CD45 APC/Cy7 (clone 30-F11; (6 citations) CD45-APC/Cy7 (clone 30-F11, (5 citations) CD45-APC/Cy7 (clone 2D1, (3 citations) APC/Cy7-conjugated anti-human CD45 (3 citations) Anti-mouse CD45-APC-Cy7 antibodies (2 citations) APC/Cy7–conjugated anti-CD45 antibody (2 citations) APC-Cy7 anti-mouse CD45 antibody (2 citations) APC/Cy7-conjugated rat anti-mouse CD45 antibody (2 citations)

107 protocols using cd45 apc cy7

Immunophenotypic Characterization of Liver Cells

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The following fluorescence-conjugated antibodies were used for KC characterization: APC Cy7 CD45 (BioLegend 103116), APC CD64 (BioLegend 139306), PE Cy7 CD11b (BioLegend, 101216), PE F4/80 (serotec, MCA497), FITC KI-67 (eBioscience 11-5698-80), Purified CD16/32 (BioLegend 101302), and the relevant isotype controls. KCs were defined as CD45⁺/CD11b^int/F4/80^hi/CD64⁺ cells^{39 (link)}.
Antibodies used for liver monocyte and neutrophil characterization included: APC Cy7 CD45 (BioLegend 103116), PE F4/80 (serotec, MCA497), PE-Cy7 CD11b (BioLegend, 101216), PerCP Cy 5.5 LY6C (BioLegend, 128012), Pacific blue MHC class II (I-Ab, BioLegend 107620), Alexa Fluor® 488 CX3CR1 (BLG-149021), Purified CD16/32 (BioLegend, 101302) and the relevant isotype controls.
Antibodies used for BM and blood monocyte characterization included: APC Cy7 CD45 (BioLegend 103116), PE-Cy7 CD11b (BioLegend, 101216), Brilliant Violet 421 LY6G (BioLegend, 127627), APC CD115 (eBioscience 17-1152-82), PerCP Cy 5.5 LY6C (BioLegend, 128012), Purified CD16/32 (BioLegend, 101302) and the relevant isotype controls.
Cells were analyzed on a Gallios^TM Flow Cytometer (Beckman Coulter) and sorted on a BD FACSAria Cell Sorter (BS Biosciences). Flow cytometry analysis was performed using Kaluza Analysis Software.

Gilboa D., Haim-Ohana Y., Deshet-Unger N., Ben-Califa N., Hiram-Bab S., Reuveni D., Zigmond E., Gassmann M., Gabet Y., Varol C, & Neumann D. (2017). Erythropoietin enhances Kupffer cell number and activity in the challenged liver. Scientific Reports, 7, 10379.

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Liver Macrophage Phenotyping and Pyroptosis

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Animals were sacrificed and livers were collected as described above. Fractionated immune cells were collected. Cells were incubated with CD16/CD32 FcR-blocking antibody (BD Bioscience, 553142) on ice for 20 minutes. Live/Dead staining was performed using Zombie Aqua (Biolegend, 423101) with a 30-minute incubation on ice. HMΦ surface staining was performed using the following panel: APC-Cy7_CD45 (BioLegend, 103116), FITC_I-A/I-E (MHCII) (BioLegend, 107605), PerCP-Cy5.5_CD11b (BioLegend, 101228), BV510_Ly6G (BioLegend, 127633), BUV395_F4/80 (BD Biosciences, 565614), BV421_CCR2 (BioLegend, 150605), APC_Ly6C (BioLegend, 128016), PE-Cy7_CD206 (ThermoFisher, 25-2061-82), BV785_CD86 (BioLegend, 105043). HMΦ pyroptosis was performed using the following panel: FAM-LEHD-FMK (caspase-11 activity assay), APC-Cy7_CD45 (BioLegend, 103116), BV510_Ly6G (BioLegend, 127633), PerCP-Cy5.5_CD11b (BioLegend, 101228), Ly-6C_AF700 (BioLegend, 128024), BUV395_F4/80 (BD Biosciences, 565614), GSDMDC1_AF674 (Santa Cruz Biotechnology, sc-393581 AF647). Flow cytometric data was acquired using LSR-II Flow Cytometer (BD Bioscience). Mean fluorescent intensity (MFI) and population percentages were analyzed using FlowJo (Ashland, OR).

Drummer C I.V., Saaoud F., Jhala N.C., Cueto R., Sun Y., Xu K., Shao Y., Lu Y., Shen H., Yang L., Zhou Y., Yu J., Wu S., Snyder N.W., Hu W., Zhuo J.‘., Zhong Y., Jiang X., Wang H, & Yang X. (2023). Caspase-11 promotes high-fat diet-induced NAFLD by increasing glycolysis, OXPHOS, and pyroptosis in macrophages. Frontiers in Immunology, 14, 1113883.

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Multiparametric Flow Cytometry for Immune Cell Profiling

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Two panels of fluorochrome-conjugated antibodies (BioLegend, San Diego, CA, USA) were used to identify the innate immunity cell populations and to analyze the activation marker expression: (1) CD11b-PE/Cy7, CD11c-PE, MHCII-Alexa488, CD103-PerCP-Cy5.5, CD45-APC/Cy7, CD64-BV421, CD24-BV510; (2) CD45-APC/Cy7, MHCII-Alexa488, Ly6G-PerCP-Cy5.5, CD86-BV421, CD83-BV510. T-cells staining was performed using the fluorochrome-conjugated antibody set containing CD4-PerCP-Cy5.5, CD8-PE/Cy7, CD62L-APC/Cy7, and CD44-BV421 (BioLegend, San Diego, CA, USA). Intracellular production of cytokines was assessed using antibodies against IFNγ-FITC, IL2-PE, and TNFα-BV510 (BioLegend, San Diego, CA, USA). Staining for the detection of intracellular markers was performed using the Fixation and Permeabilization Solution reagent kit (BD Biosciences, San Jose, CA, USA) according to the manufacturer’s instructions. Zombie Red viability marker (BioLegend, San Diego, CA, USA) was used to identify the dead cells. True Stain reagent (BioLegend, San Diego, CA, USA), containing antibodies to CD16/CD32, was added during the surface markers staining to block non-specific antibody binding. Data were collected on a Cytoflex flow cytometer (Beckman Coulter, Bray, CA, USA). The results were analyzed using the Kaluza Analysis 2.2 program (Beckman Coulter, Bray, CA, USA).

Vasilyev K., Shurygina A.P., Zabolotnykh N., Sergeeva M., Romanovskaya-Romanko E., Pulkina A., Buzitskaya J., Dogonadze M.Z., Vinogradova T.I, & Stukova M.A. (2021). Enhancement of the Local CD8+ T-Cellular Immune Response to Mycobacterium tuberculosis in BCG-Primed Mice after Intranasal Administration of Influenza Vector Vaccine Carrying TB10.4 and HspX Antigens. Vaccines, 9(11), 1273.

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Characterization of Murine Skin Immune Cells

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Skin from mice was isolated and subjected to mild digestion with liberase (Roche™), and mechanically disrupted using GentleMACS Dissociator (Miltenyi). The single cell suspension was stained with either CD45-APCCy7 (Biolegend), CD3-PE (Biolegend), gdTCR-Brilliant Violet 421 (Biolegend), CD90.2-APC (BD PharMingen), Cd11b-PerCPCy5.5 (BD PharMingen) and CD2-PECy7 (Biolegend), or CD45-APCCy7 (Biolegend), MHCII-PacBlue (Biolegend), Ly6C-APC (BD PharMingen), Ly6G-PE (BD PharMingen), CD11b-PECy7 (BD PharMingen). Samples were collected in a FACS CANTO II (BD, San Jose CA) equipped with 488nm, 640nm and 405nm lines. We used pulse processing to exclude cell aggregates and live/dead fixable dye Aqua (Invitrogen) to exclude dead cells. At least 100,000 alive single events were collected; all data were analyzed using FlowJo v10 (Treestar, Oregon).

Uluçkan Ö., Jiménez M., Roediger B., Schnabl J., Díez-Córdova L.T., Troulé K., Weninger W, & Wagner E.F. (2019). Cutaneous Immune Cell-Microbiota Interactions Are Controlled by Epidermal JunB/AP-1. Cell Reports, 29(4), 844-859.e3.

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Multicolor Flow Cytometry Analysis of Tumor-Infiltrating Lymphocytes

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Tumors were incubated with collagenase/DNase for 15 min, homogenized and first incubated with Fc Block™ (BD-Biosciences) for 10 min. Then, tumor samples were stained with the following antibodies to simultaneously quantify lymphocyte subsets, OVA(257-264) specific CD8 T-cells and expression of immune checkpoint molecules: CD45-APC-Cy7, K^b/OVA(257-264)-APC, CD4-Alexa 700, CD8-PE, CD3-BUV496, PD1-PerCP/Cy5.5, LAG3-BV650 and TIM3-BV785 (all from Biolegend, San Diego, CA, USA, except CD3-BUV496 which is from BD-Biosciences).
To analyze effector functions cells were stimulated for 4 h with PMA (100 ng/mL)/ionomycin (1 μg/mL) and then stained with antibodies against CD45-APC-Cy7, CD4-Alexa 700, CD8-BV421 CD107a-FITC (all from Biolegend), CD3-BUV496 (from BD-Biosciences). Then cells were fixed and permeabilized and finally stained intracellularly with anti-IFN-γ-PE and TNF-α-BV510.
In all cases the promofluor 840 (maleimide, Promokine, Burlingame, CA, USA) was added to stain dead cells. Samples were acquired with FACSCantoII (Becton Dickinson, Franklin Lakes, NJ, USA) or with Cytoflex (Beckman Coulter, Irving, TX, USA) flow cytometers and analyzed using FlowJo software (Tree Star Inc., Ashland, OR, USA).

Silva L., Egea J., Villanueva L., Ruiz M., Llopiz D., Repáraz D., Aparicio B., Lasarte-Cia A., Lasarte J.J., Ruiz de Galarreta M., Lujambio A., Sangro B, & Sarobe P. (2020). Cold-Inducible RNA Binding Protein as a Vaccination Platform to Enhance Immunotherapeutic Responses against Hepatocellular Carcinoma. Cancers, 12(11), 3397.

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Multiparametric Flow Cytometry Analysis

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PE-Cy™7-CD4 (Clone: RM4-5), APC-Cy™7-CD69 (Clone: H1.2F3), APC-Cy™7-CD45 (Clone: 30-F11), APC-Cy™7-CD19 (Clone: 6D5), PE-Cy™7-MHC II (Clone: M5/114.15.2), PE-Foxp3 (Clone: MF-14), FITC-Ly6C (Clone: HK1.4), BV421-CD11b (Clone: M1/70) were purchased from Biolegend. PE-F4/80 (Clone: BM8), APC-IFN-γ (Clone: XMG1.2), Biotin-CD11c (Clone: N418) were purchased from eBioscience. APC-Cy™7-CD4 (Clone: GK1.5), APC-CD40 (Clone: HM40-3), APC-CD25 (Clone PC61), PerCP-Cy™5.5-CD80 (Clone: 16-10A1), FITC-CD3e (Clone: 145-2C11), PE-IL-17a (Clone: TC11-18H10), Sav-BUV395 were purchased from BD Biosciences. Flow cytometry data were acquired on BD LSR II flow cytometer as previously described⁴⁰ and analyzed with FlowJo™ v10.7.

Guan D., Wang Z., Huo J., Xu S, & Lam K.P. (2021). Bruton’s tyrosine kinase regulates gut immune homeostasis through attenuating Th1 response. Cell Death & Disease, 12(5), 431.

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Isolation and Flow Cytometry of Murine Splenic Dendritic Cells

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The spleen was dissociated into a single‐cell suspension using PBS supplemented with 10% fetal bovine serum in GentleMACs tubes (Miltenyi Biotec). Red blood cells were lysed by resuspending splenic cells in ammonium chloride‐Tris buffer and incubation at room temperature for 8 minutes. Leukocytes were collected following centrifugation and washed twice in PBS. The splenic leukocytes were resuspended in PBS and enriched to 2×10⁶/μL. LIVE/DEAD Fixable Aqua Dead Cell Stain Kit (ThermoFisher) was used to check the viability of splenic leukocytes. Specimens (100 μL each) were treated with CD16/32 blocking antibodies for 10 minutes followed by antibody mixtures (APC/Cy7‐CD45, FITC‐CD3, PerCP/Cy5.5‐B220, APC‐IA/IE, PE‐PDCA1, BV421‐CD11c) (BioLegend, San Diego CA) for 30 minutes. Corresponding fluorescence minus one controls were also prepared. Then, all specimens were fixed by adding 500 μL 4% paraformaldehyde (Sigma‐Aldrich). Flow cytometry was performed using Attune NxT Flow Cytometer (ThermoFisher), and data were analyzed using FlowJo software (BD Company, Ashland, OR). The pDCs were defined by CD45⁺, CD3‐/IA/IE⁺ and B220+/PDCA1⁺, and cDCs by CD45⁺ and CD11c+/IA/IE⁺.

Lai L., Zhang A., Yang B., Charles E.J., Kron I.L, & Yang Z. (2021). Plasmacytoid Dendritic Cells Mediate Myocardial Ischemia/Reperfusion Injury by Secreting Type I Interferons. Journal of the American Heart Association: Cardiovascular and Cerebrovascular Disease, 10(15), e020754.

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Fluorescent Labeling and Cell Staining

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Human serum albumin (HSA), 2-(N-Morpholino) ethanesulfonic acid (MES), fluorescein isothiocyanate (FITC) were purchased from Sigma-Aldrich (Shanghai) Trading Co., Ltd. MTX (≥98%), sodium dodecyl sulfate (SDS, ≥98%), dimethylsulfoxide (DMSO, ≥99%) were obtained from Shanghai Aladdin Biochemical Technology Co., Ltd. Sodium hyaluronate (5 kDa) was purchased from Bloomage Freda Biopharm Co., Ltd., (Jinan, China). Paraformaldehyde (4%) was obtained from Beyotime Biotechnology Co., Ltd. Imiquimod cream (5%) was from Mingxin Pharmaceuticals (Sichuan, China). CD16/32, APC-Cy7-CD45, FITC-CD3, Qdot-605-CD11c were bought from BioLegend Inc. RPMI 1640 medium and fetal bovine serum (FBS) were purchased from Thermo Fisher Scientific Inc.

Wang H., Zhao Z., Wu C., Tong X., Shi Y, & Chen S. (2022). Microneedle Patch Delivery of Methotrexate-Loaded Albumin Nanoparticles to Immune Cells Achieves a Potent Antipsoriatic Effect. International Journal of Nanomedicine, 17, 3841-3851.

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Comprehensive Cell Line Validation and Inhibitor Characterization

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All cell lines were obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA), independently validated by short tandem repeat (STR) DNA fingerprinting, and tested negative for mycoplasma infection. NCI-H157, A549, NCI-H1299, HeLa, BT549, MDA-MB-231, CT26, B16-F10, MC-38, LLC and 293T cells were cultured in RPMI 1640 or DMEM media, supplemented with 10% fetal bovine serum (Thermo Fisher Scientific, Waltham, MA, USA) at 37 °C in a humidified atmosphere with 5% CO₂. Inhibitors for ATM (KU-60019, AZD1390, AZD0156), STING (H151), p-TBK1 (GSK8612), p-STAT1 (fludarabine) were purchased from Selleck (Houston, TX, USA). Human IFNβ and Galectin-9 ELISA assay Kits were purchased from R&D SYSTEMS (Minneapolis, MN, USA). Anti-human Gal-9 antibody was purchased from BioRad (Hercules, CA, USA). Anti-mouse Gal-9 antibody and IgG isotype control were purchased from BioXCell (Lebanon, NH, USA). Antibodies against cGAS, STING, p-STING(Ser366), p-TKB1(Ser172), TKB1, p-STAT1(Tyr701), STAT1 and Actin were purchased from Cell Signaling Technology (Cambridge, MA, USA). Anti-mouse fluorochrome-conjugated antibodies including FITC-CD4, Percp/Cy5.5-CD8, AF700-CD3, APC/Cy7-CD45, PE/Cy7-IFNγ antibodies were purchased from BioLegend (San Diego, CA, USA).

Zheng S., Song J., Linghu D., Yang R., Liu B., Xue Z., Chen Q., Liu C., Zhong D., Hung M.C, & Sun L. (2023). Galectin-9 blockade synergizes with ATM inhibition to induce potent anti-tumor immunity. International Journal of Biological Sciences, 19(3), 981-993.

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Immunophenotyping of Cell Populations

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A total of 3 × 10⁵ CMSCs per line were used for staining and flow cytometry analysis. Samples were incubated with primary antibodies at room temperature for 15 min. For indirect staining, cells were washed in FACS media (PBS + FBS 2%) after the first incubation, centrifuged for 5 min, resuspended with the secondary antibody, and incubated at room temperature for 20 min. The antibodies used were: anti-DDR2 (sc-81707, Santa Cruz Biotechnology, Inc., Dallas, TX, USA) + goat anti-mouse Alexa Fluor 647 (Invitrogen); PerCP/Cy5.5-CD90 (202515, BioLegend, San Diego, CA, USA); APC/Cy7-CD45 (202216, BioLegend, San Diego, CA, USA); and APC-CD31 (FAB3628A, R&D systems, Minneapolis, MN, USA). Sample acquisition was performed on a FACSAria™ II flow cytometer equipped with the 6.1.1 Diva software (BD Biosciences, Franklin lakes, NJ, USA). Data analysis was performed using FlowJo software, version 10.6.1.

Picchio V., Gaetani R., Pagano F., Derevyanchuk Y., Pagliarosi O., Floris E., Cozzolino C., Bernava G., Bordin A., Rocha F., Pereira A.R., Ministro A., Pinto A.T., De Falco E., Serino G., Massai D., Tamarat R., Pesce M., Santos S.C., Messina E, & Chimenti I. (2024). Early Impairment of Paracrine and Phenotypic Features in Resident Cardiac Mesenchymal Stromal Cells after Thoracic Radiotherapy. International Journal of Molecular Sciences, 25(5), 2873.

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