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Apc anti mouse cd11c

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APC anti-mouse CD11c is a fluorochrome-conjugated antibody that binds to the CD11c antigen expressed on the surface of mouse dendritic cells. It is used in flow cytometry applications to identify and quantify dendritic cell populations in mouse samples.

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Lab products found in correlation

Anti fcγ receptor, by BD (2 mentions) Pe anti mouse cd3ε, by BioLegend (2 mentions) Apc cy7 anti mouse f4 80, by BioLegend (2 mentions) Fitc conjugated anti mouse ifn α, by PBL Assay Science (2 mentions) Anti mouse cd11b pe cy7, by Thermo Fisher Scientific (2 mentions) Anti mouse cd45 efluor450, by Thermo Fisher Scientific (2 mentions) Facscalibur cytometer, by BD (1 mentions) Percp cy5.5 anti mouse cd45r b220, by BD (1 mentions) Lsrfortessa, by BD (1 mentions) Cell dissociation buffer, by Thermo Fisher Scientific (1 mentions) Live dead fixable violet dead cell stain, by Thermo Fisher Scientific (1 mentions) Chloroquine, by Merck Group (1 mentions) Ca 074 me, by Merck Group (1 mentions) Flt3l, by R&D Systems (1 mentions) Bafilomycin a1, by Merck Group (1 mentions) Anti mouse cd3e, by BioLegend (1 mentions) Kaluza software, by Beckman Coulter (1 mentions) 70 m cell strainer, by BD (1 mentions) Cytofix fixation buffer, by BD (1 mentions) Perm buffer 3, by BD (1 mentions)

8 protocols using apc anti mouse cd11c

1

Multiparameter Flow Cytometry Analysis of Mammary Gland

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Mammary glands and tumors were digested and brought to single cell suspension as described above and with the presence of Golgi-plug as indicated by the manufacturer (BD Biosciences). After fixation with 2% paraformaldehyde 30 minutes on ice, cells were permeabilized and blocked with 0.2% saponin, 5 mM EDTA, 2 mM NaN3, 5% NGS and 4 μg/ml anti-Fcγ receptor (Clone 2.4 G2; BD Biosciences). IFN-α was stained using FITC-conjugated anti-mouse IFN-α (Clone RMMA-1; PBL)67 (link). T-cells were stained with PE anti-mouse CD3ε (Clone 145-2C11; BioLegend), macrophages with APC/Cy7 anti-mouse F4/80 (Clone BM8; BioLegend), and dendritic cells with APC anti-mouse CD11c (Clone HL3; BD Pharmingen). FITC-conjugated rat IgG1 was used as negative isotype control (Clone RTK2071; BioLegend). Note: pregnancy, lactation and involution cell samples display small levels of auto-fluorescence for reasons unknown. We thoroughly excluded this population of our analysis and applied the same gating for all the all samples.
Celià-Terrassa T., Liu D., Choudhury A., Hang X., Wei Y., Zamalloa J., Alfaro-Aco R., Chakrabarti R., Jiang Y.Z., Koh B.I., Smith H., DeCoste C., Li J.J., Shao Z.M, & Kang Y. (2017). Normal and cancer mammary stem cells evade interferon-induced constraint through the miR-199a-LCOR Axis. Nature cell biology, 19(6), 711-723.
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2

Assessing BMDC Maturation Status

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At the end of the stimulation, the maturation status of BMDCs was assessed. In brief, the co-cultured and undisposed BMDCs were gathered, rinsed twice with PBS, and three fold-labeled for half an hour at 4°C, protected from light, with APC-anti-mouse CD11c, FITC-anti-mouse CD80, and PE-anti-mouse CD86 monoclonal antibodies (BD Biosciences) as DC maturation markers. The corresponding isotype control antibodies were used as controls. After thorough rinsing by washing twice with PBS, expression levels of associated surface molecules in labeled BMDCs were determined by flow cytometry in a FACS Calibur cytometer (BD Biosciences). The data were analyzed using FlowJo software (version 7.6).
Zheng S., Qin T., Lu Y., Huang Y., Luo L., Liu Z., Bo R., Hu Y., Liu J, & Wang D. (2018). Maturation of dendritic cells in vitro and immunological enhancement of mice in vivo by pachyman- and/or OVA-encapsulated poly(d,l-lactic acid) nanospheres. International Journal of Nanomedicine, 13, 569-583.
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3

Multiparameter Flow Cytometry Analysis of Mammary Gland

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Mammary glands and tumors were digested and brought to single cell suspension as described above and with the presence of Golgi-plug as indicated by the manufacturer (BD Biosciences). After fixation with 2% paraformaldehyde 30 minutes on ice, cells were permeabilized and blocked with 0.2% saponin, 5 mM EDTA, 2 mM NaN3, 5% NGS and 4 μg/ml anti-Fcγ receptor (Clone 2.4 G2; BD Biosciences). IFN-α was stained using FITC-conjugated anti-mouse IFN-α (Clone RMMA-1; PBL)67 (link). T-cells were stained with PE anti-mouse CD3ε (Clone 145-2C11; BioLegend), macrophages with APC/Cy7 anti-mouse F4/80 (Clone BM8; BioLegend), and dendritic cells with APC anti-mouse CD11c (Clone HL3; BD Pharmingen). FITC-conjugated rat IgG1 was used as negative isotype control (Clone RTK2071; BioLegend). Note: pregnancy, lactation and involution cell samples display small levels of auto-fluorescence for reasons unknown. We thoroughly excluded this population of our analysis and applied the same gating for all the all samples.
Celià-Terrassa T., Liu D., Choudhury A., Hang X., Wei Y., Zamalloa J., Alfaro-Aco R., Chakrabarti R., Jiang Y.Z., Koh B.I., Smith H., DeCoste C., Li J.J., Shao Z.M, & Kang Y. (2017). Normal and cancer mammary stem cells evade interferon-induced constraint through the miR-199a-LCOR Axis. Nature cell biology, 19(6), 711-723.
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4

Quantifying Stress-Induced Translational Regulation

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Total splenocytes isolated from HF or DMSO pre-conditioned mice were surface stained using PerCP Cy5.5 anti-mouse CD45R/B220 (BD Pharmingen), APC anti-mouse CD11c (BD Pharmingen), and Alexa-Fluor 700 anti-mouse CD3 (eBioscience) at 4°C for 30 min. The stained cells were washed with FACS buffer and quickly fixed using 4% paraformaldehyde followed by intracellular staining for eIF2α-P as described earlier (33 ). Fixed cells were permeabilized using 1X Perm Wash Buffer (Bio Legend) and incubated with rabbit anti-mouse monoclonal eIF2α-P antibody (Cell Signalling) or respective isotype control overnight at 4°C. Following washes with 1X FACS buffer, the cells were stained with anti-mouse Alexa488 secondary antibody for 1h at room temperature, washed and re-suspended in FACS buffer and acquired on BD LSR Fortessa (BD Biosciences) cytometer and analyzed using FlowJo (Tree Star Inc).
Afroz S., S.h.a.m.a., Battu S., Matin S., Solouki S., Elmore J.P., Minhas G., Huang W., August A, & Khan N. (2019). Amino acid starvation enhances vaccine efficacy by augmenting neutralizing antibody production. Science signaling, 12(607), eaav4717.
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5

Investigating CD11c+ Dendritic Cell Activation

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After CD11c+ cell isolation, cells were seeded in 24-well plates at varying ratios of cells from the CD11c enriched isolate to cells from the CD11c reduced isolate. Ratios of 8:1, 1:1, and 1:8 were investigated. Wells with only CD11c enriched and reduced isolate cells were included as controls. Total cell number was kept constant for all ratios at 250,000 cells/well. After seeding, cells received either media only or stimulation with MPLA (10 μg/mL) (InvivoGen, San Diego, CA). Lipopolysaccharide (LPS) stimulation (1 μg/mL) of CD11c enriched cells was included in the first experiment as a positive control. Twenty-four hours later, all cells were collected using Cell Dissociation Buffer (Thermo Fisher Scientific, Waltham, MA), blocked with anti-Fc antibody, and stained with APC anti-mouse CD11c, PerCP/Cy5.5 anti-mouse MHC Class II (MHCII), Brilliant Violet (BV) 605 anti-mouse CD86 (antibodies purchased from BD Biosciences, Franklin Lakes, NJ), and LIVE/DEAD® Fixable Violet Dead Cell Stain (Thermo Fisher Scientific). After fixation with 1.6% paraformaldehyde, cells were analyzed by flow cytometry (FACSCanto II). Flow cytometry data were analyzed by FlowJo software for surface marker expression of the live, CD11c+ gated population.
Frizzell H., Park J., Lou N.C, & Woodrow K.A. (2016). Role of heterogeneous cell population on modulation of dendritic cell phenotype and activation of CD8 T cells for use in cell-based immunotherapies. Cellular immunology, 311, 54-62.
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6

Dendritic Cell Activation Assay

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The commercial sources for reagents were as follows: CpG oligodeoxynucleotide ODN2216 (Invivogen); chloroquine, bafilomycin A1 and CA-074-Me (Sigma-Aldrich); Flt3L (R & D systems); anti-mouse CD11c APC (BD Pharmingen), anti-mouse B220 APC-Cy7 and anti-mouse PDCA-1 PE (Milteny Biotec); and Phosphonoacetate (Sigma).
Dai P., Wang W., Cao H., Avogadri F., Dai L., Drexler I., Joyce J.A., Li X.D., Chen Z., Merghoub T., Shuman S, & Deng L. (2014). Modified Vaccinia Virus Ankara Triggers Type I IFN Production in Murine Conventional Dendritic Cells via a cGAS/STING-Mediated Cytosolic DNA-Sensing Pathway. PLoS Pathogens, 10(4), e1003989.
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7

Comprehensive Murine Immune Cell Analysis

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Mouse ears were subjected to enzymatic digestion as previously described (Feuerstein et al., 2015 (link)). After digestion, samples were filtered with a 70 µm cell strainer (BD), washed with FACS Buffer (PBS+2%FBS+2 mM EDTA) and stained with the indicated antibodies. The following antibodies were used: anti-mouse CD45 eFluor450 (eBioscience), anti-mouse CD11b PE-Cy7 (eBioscience), anti-mouse Ly6G FITC (BD Biosciences), anti-mouse Ly6C PerCP-Cy5.5 (BD Biosciences), anti-mouse CD64 PerCP/Cy5.5 (Biolegend), anti-mouse CD11c APC (BD Biosciences), mouse MHC class II eFluor450 (eBiosciences) and anti-mouse CD3e (Biolegend). Cell samples were analyzed with a 10-color flow cytometer (Gallios, Beckman Coulter) and the Kaluza software (version 1.5a, Beckman Coulter).
Feuerstein R., Forde A.J., Lohrmann F., Kolter J., Ramirez N.J., Zimmermann J., Gomez de Agüero M, & Henneke P. (2020). Resident macrophages acquire innate immune memory in staphylococcal skin infection. eLife, 9, e55602.
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8

Phosphorylation of Stat1 in Immune Cells

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Mouse ears were subjected to enzymatic digestion, filtered, washed and cells finally resuspended in 100 µl RPMI medium supplemented with 10% FBS and antibiotics (ciprofloxacin, 10 mg/ml). Then cells were stimulated for 15 min at 37°C with 200 ng/ml IFNγ and subsequently fixed with 100 µl fixation buffer (Cytofix Fixation Buffer, BD Biosciences) for 10 min at 37°C. After centrifugation and permeabilization with 400 µl ice cold Perm Buffer III (BD Phosflow) for 30 min at 4 °C cells were washed twice with PBS containing 1% BSA. Then cell surfaces were stained with anti-mouse CD45 eFluor450 (eBioscience), anti-mouse CD11b PE-Cy7 (eBioscience), anti-mouse CD11c APC (BD Biosciences) and intracellular phosphorylated Stat1 stained with mouse anti-Stat1 (pY701) Alexa Fluor 647 (BD Biosciences) 30 min at RT and subsequently analyzed by flow cytometry.
Feuerstein R., Forde A.J., Lohrmann F., Kolter J., Ramirez N.J., Zimmermann J., Gomez de Agüero M, & Henneke P. (2020). Resident macrophages acquire innate immune memory in staphylococcal skin infection. eLife, 9, e55602.
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