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Cd11b apc cy7

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CD11b-APC-Cy7 is a fluorophore-conjugated antibody that binds to the CD11b surface antigen. CD11b is a subunit of the integrin receptor Mac-1 (CD11b/CD18), which is expressed on the surface of various immune cells such as monocytes, macrophages, and granulocytes. The APC-Cy7 fluorescent label allows for the detection and analysis of CD11b-expressing cells using flow cytometry.

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Spelling variants of this product

CD11b (457 citations) CD11b-APC (115 citations) APC-Cy7 (106 citations) Anti-CD11b-APC-Cy7 (26 citations) APC-Cy7 anti-mouse CD11b (5 citations) APC/Cy7-conjugated anti-mouse CD11b (3 citations) B220-APC and CD11b-APC/cy7 antibodies (3 citations) Anti-F4/80 Alexa Fluor-488 and anti-CD11b APC/Cy7 (2 citations) APC-Cy7-α-CD11b (M1/70, (2 citations) Anti-CD11b APC-Cy7-conjugated antibody (2 citations)

38 protocols using cd11b apc cy7

1

Immune Cell Profiling by Flow Cytometry

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Antibodies used in the studies are mentioned below:
Anti-mouse: CD3-Pacific Blue, CD4-PE, CD8-APCCy7, CD69-FITC, CD44-FITC, CD62L-APC, IFNγ-APC, IL-17-PECy7, CD11b-APCCy7, CD11c-APC, CD80-FITC, CD86-PerCPCy5.5, CD40-PE, CD4-APC, and CD4-FITC from Biolegend, USA.
Anti-mouse: p38, ph-p38 and β-Actin from Cell Signaling Technology.
Pahuja I., Verma A., Ghoshal A., Mukhopadhyay S., Kumari A., Shaji A., Chaturvedi S., Dwivedi V.P, & Bhaskar A. (2023). Biapenem, a Carbapenem Antibiotic, Elicits Mycobacteria Specific Immune Responses and Reduces the Recurrence of Tuberculosis. Microbiology Spectrum, 11(4), e00858-23.
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2

Comprehensive Flow Cytometry and Proliferation Assays

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Flow cytometry was performed on dissociated MCT or HEK293T cells stained with the Fn14-APC antibody (1:30; Miltenyi, 130-104-281). For negative control, unstained cells or the respective isotype control antibody, mouse IgG2b (1:100, Miltenyi, 130-098-890) was used. Antibody staining was conducted for 30 min in the dark at 4°C. Proliferation assay was conducted as follows: from naïve animal’s lymph node/Spleen, T cells were isolated and stained with 1:1000 e450 proliferation dye (ThermoFisher, 65-0842-85) and treated with various reagents (11R-VIVIT, CsA). Cells were then stimulated with ⍺CD3/⍺CD28 dynabeads for 5 days. White blood cells were stained for 30 min in the dark at 4°C with CD45-PerCP/Cy5-5 (1:100; Biolegend, 103132) and CD11b-APC/Cy7 (1:100; Biolegend, 101226). The kidney cell suspension was stained with CD45-PerCP/Cy5-5 and CD326-PE (1:50, Miltenyi, 130-117-667) for 30min in the dark at 4°C. Also, single stains were created to perform proper compensation.
Karolin A., Escher G., Rudloff S, & Sidler D. (2022). Nephrotoxicity of Calcineurin Inhibitors in Kidney Epithelial Cells is Independent of NFAT Signaling. Frontiers in Pharmacology, 12, 789080.
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3

Comprehensive Immune Cell Analysis

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The following antibodies were used to characterize BMDCs and assess their activation state: MHC II-PE, CD11c-APC, CD86-PE-Cy7, CD80-FITC and CD11b-APC-Cy7 (all from BioLegend). Acquisition was performed on an Attune NxT (ThermoFisher). Leukocyte populations from BM, liver and spleen were also analyzed by flow cytometry. Single cells were excluded from dead cells using the LIVE/ DEAD Zombie NIR Fixable Viability Kit (BioLegend). Immuno-phenotyping was performed using the following antibodies: CD45-BV510, CD49b-PE-Dazzle, CD19-PerCP, CD3-FITC, CD4-AF700, CD8-BV785, C44-BV650 and CD62L-BV421 (all from BioLegend). Full minus one (FMO) controls were used to determine positivity. Precision count beads (BioLegend) were used to count immune cells in different organs. Before acquisition, stained cells were fixed with 1% Paraformaldehyde (Sigma-Aldrich). Acquisition was performed using the BD LSRFortessa and data were analyzed using the FlowJo software (TreeStar) version 10.
Helou D.G., Mauras A., Fasquelle F., Lanza J.S., Loiseau P.M., Betbeder D, & Cojean S. (2021). Intranasal vaccine from whole Leishmania donovani antigens provides protection and induces specific immune response against visceral leishmaniasis. PLoS Neglected Tropical Diseases, 15(8), e0009627.
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4

Flow Cytometry Analysis of Immune Cell Subsets

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Skin samples were prepared as described previously [15 (link)], and flow cytometry was performed as described previously [16 (link)]. In brief, single-cell suspensions (1 million cells) were first incubated with Fc Block (BD Pharmingen, San Diego, CA) for 10 minutes, then coincubated with antibodies for 20 minutes at 4 °C, followed by washing with staining buffer (PBS + 1% FBS). Flow cytometry analysis was performed with an LSRFortessa cell analyzer (BD Biosciences, San Jose. CA), and data were analyzed using Flowjo software (Flowjo, Ashland, OR). CD19-FITC (BD Pharmingen), CD229.1-PE (BD Pharmingen), CD4-PE-Cy7 (eBioscience, San Diego, CA), CD25-APC-Cy7 (BD Biosciences), CD45-PB (BioLegend San Diego, CA), CD8α-AF700 (BioLegend), B220-PE (BD Biosciences), CD138-PE (BioLegend). CD11c-FITC (BioLegend). F4/80-APC (eBioscience), and CD11 b-APC-Cy7 (BioLegend) were used for these studies.
Pai C.C., Khuat L.T., Chen M., Murphy W.J, & Abedi M. (2016). Therapeutic Effects of a NEDD8-Activating Enzyme Inhibitor, Pevonedistat, on Sclerodermatous Graft-versus-Host Disease in Mice. Biology of blood and marrow transplantation : journal of the American Society for Blood and Marrow Transplantation, 23(1), 30-37.
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5

Multiparametric Immunostaining of Dissociated Cells

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Dissociated cells were briefly fixed in 4% paraformaldehyde for 15 min at room temperature and washed once with FACS staining buffer (5% fetal bovine serum (FBS) in PBS). Cells were then resuspended in 100 μl of saponin buffer (0.5% saponin and 2% FBS in PBS) and immunostained with anti-CD115-PE cy7 (1:1000, eBioscience #25115282), CD11b-APC-cy7 (1:2000, Biolegend #101226), PU.1 (1:1000, Cell Signaling #2266), chicken anti-GFP antibody (1:1000, Abcam #ab13970), anti-CX3CR1-APC (1:2000, Biolegend #149008), or anti-Ki67-PerCP-efluro710 (1:2000, Invitrogen, #66–5698–82) for 60 min at 4 °C57 (link).
Yu X., Liu H., Hamel K.A., Morvan M.G., Yu S., Leff J., Guan Z., Braz J.M, & Basbaum A.I. (2020). Dorsal root ganglion macrophages contribute to both the initiation and persistence of neuropathic pain. Nature Communications, 11, 264.
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6

Immune Cell Profiling of Islets

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Islets were isolated from ZBTB46-DTR BM chimera mice dissociated as previously described. Islet samples were stained with the fluorescent-labeled antibodies CD45 BUV395 (BD), CD90.2 PE-Cy7 (BioLegend), CD19 PE (BioLegend), F4/80 PerCP-Cy5.5 (BioLegend), MHC-II FITC (BioLegend), CD11c BUV805 (BD), CD11b APC-Cy7 (BioLegend), XCR1 BV785 (BioLegend), unlabeled goat α-MERTK (R&D Systems), and donkey α-goat secondary (AF647; Jackson ImmunoResearch).
Lindsay R.S., Whitesell J.C., Dew K.E., Rodriguez E., Sandor A.M., Tracy D., Yannacone S.F., Basta B.N., Jacobelli J, & Friedman R.S. (2021). MERTK on mononuclear phagocytes regulates T cell antigen recognition at autoimmune and tumor sites. The Journal of Experimental Medicine, 218(10), e20200464.
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7

Multiparameter Flow Cytometry Profiling

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Cells were dissociated with Accutase for 20min and resuspended in FACS buffer containing 1% BSA, 2mM EDTA, 30ug/mL DNAse I and Normocin in PBS. Cells were washed and incubated in FACS buffer with antibody for 30 minutes on ice at 4 degrees in the dark. Cells were washed and resuspended in FACS buffer and strained through 40uM caps to eliminate cell clumps. Data collection was done on a BD-LSRII and FACSDiva v8. Gating (Fig. S2) and subsequent analysis was done using FlowJo software v10.5.3. FACS antibodies used include KDR-PE – R&D, FAB357P, Clone 89106 at 1:60, CD235A-APC – BD Biosciences at 1:100, 551336, Clone GA-R2, CD41-Apc/Cy7 – Biolegend, 303715, Clone HIP8, CD43-PerCP/Cy5.5 -BD Biosciences, 563521, Clone 1G10, CD45-FITC – BD Biosciences, 560976, Clone HI3O, CX3CR1-PE – Biolegend, 341604, 2A9–1, and Cd11b-APC/Cy7 – Biolegend, 301351, ICRF44 all at 5ul/100ul test.
Guttikonda S.R., Sikkema L., Tchieu J., Saurat N., Walsh R., Harschnitz O., Ciceri G., Sneeboer M., Mazutis L., Setty M., Zumbo P., Betel D., de Witte L.D., Pe’er D, & Studer L. (2021). Fully defined human pluripotent stem cell-derived microglia and tri-culture system model C3 production in Alzheimer’s disease. Nature neuroscience, 24(3), 343-354.
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8

Comprehensive Immune Cell Profiling

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We used the following antibodies: CD4-PErCP (100538), CD8-APC/Cy7 (100713), CD69-FITC (104506), CD25-APC (101910), IFNγ-APC (505810), IL17-PE/Cy7 (506922), CD11b-APC/Cy7 (101226), CD11c-APC (117310), CD80-FITC (104706), MHCII-PE (107607), CD3-PacificBlue (100214), CD4-PE (100512), CD4-APC (100516) and CD4-FITC (100510) from Biolegend, USA.
SIRT2 (ab211033), β-actin (ab8227), acH3K18 (ab40888), α-tubulin (ab184613), acetyl-α-tubulin (ab179484), acetyl-NFκB p65 (ab19870), Alexa Fluor 647 (ab150075) and IgG-APC isotype control (ab232814) from Abcam.
ERK1/2 (9102), Phospho-ERK1/2 (9101), p-38 (9212) and phospho-p38 (4511) from Cell Signaling Technology.
Bhaskar A., Kumar S., Khan M.Z., Singh A., Dwivedi V.P, & Nandicoori V.K. (2020). Host sirtuin 2 as an immunotherapeutic target against tuberculosis. eLife, 9, e55415.
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9

Multiparameter Flow Cytometry Profiling

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Cells were dissociated with Accutase for 20min and resuspended in FACS buffer containing 1% BSA, 2mM EDTA, 30ug/mL DNAse I and Normocin in PBS. Cells were washed and incubated in FACS buffer with antibody for 30 minutes on ice at 4 degrees in the dark. Cells were washed and resuspended in FACS buffer and strained through 40uM caps to eliminate cell clumps. Data collection was done on a BD-LSRII and FACSDiva v8. Gating (Fig. S2) and subsequent analysis was done using FlowJo software v10.5.3. FACS antibodies used include KDR-PE – R&D, FAB357P, Clone 89106 at 1:60, CD235A-APC – BD Biosciences at 1:100, 551336, Clone GA-R2, CD41-Apc/Cy7 – Biolegend, 303715, Clone HIP8, CD43-PerCP/Cy5.5 -BD Biosciences, 563521, Clone 1G10, CD45-FITC – BD Biosciences, 560976, Clone HI3O, CX3CR1-PE – Biolegend, 341604, 2A9–1, and Cd11b-APC/Cy7 – Biolegend, 301351, ICRF44 all at 5ul/100ul test.
Guttikonda S.R., Sikkema L., Tchieu J., Saurat N., Walsh R., Harschnitz O., Ciceri G., Sneeboer M., Mazutis L., Setty M., Zumbo P., Betel D., de Witte L.D., Pe’er D, & Studer L. (2021). Fully defined human pluripotent stem cell-derived microglia and tri-culture system model C3 production in Alzheimer’s disease. Nature neuroscience, 24(3), 343-354.
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10

Intestinal Tissue-Resident Macrophage Analysis

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For flow cytometric analysis of intestinal TRMs in the lamina propria, small intestine segments were washed, flattened, cut into 1.5cm pieces and shaken in 2 changes of HBSS with 5% FBS and 2 mM EDTA for 20 min at 37C. The suspension was passed through a mesh strainer to remove epithelial cells. The intestinal pieces were chopped using scissors and shaken in digestion media containing HBSS (Gibco) with 5% FBS, 1mg/ml Collagenase D, 2U/ml DNase I, 0.1U/ml Dispase for 30 mins at 37C. The digested samples were then vortexed briefly and passed through a 100-micron filter followed by staining for flow cytometry.
Cells were stained with the following fluorochrome-conjugated surface antibodies: CD45-BUV395 (BD Biosciences; Clone 30-F11), CD64-PE/Cy7 (Biolegend; Clone X54-5/7.1), CD11c-Alexa Fluor 488 (eBioscience; Clone N418), CD11b-APC/Cy7 (Biolegend; Clone M1/70), CX3CR1-BV785 (Biolegend; Clone SA011F11), and F4/80-APC (eBioscience; Clone BM8). Cellular fluorescence was measured using an LSRII Fortessa flow cytometer, and data were analyzed using FlowJo software (Tree Star).
Martin A.T., Giri S., Safronova A., Eliseeva S.I., Kwok S.F, & Yarovinsky F. (2024). Parasite-induced IFN-γ regulates host defense via CD115 and mTOR-dependent mechanism of tissue-resident macrophage death. PLOS Pathogens, 20(2), e1011502.
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