Cd11c bv605

Manufactured by BD

CD11c-BV605 is a fluorochrome-conjugated antibody that is used to detect and analyze CD11c-expressing cells in flow cytometry applications. CD11c is a cell surface integrin that is expressed on dendritic cells, macrophages, and other myeloid cells.

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3 protocols using cd11c bv605

Characterization of Dsg3-specific B Cells

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PBMCs were obtained from blood samples of patients using Ficoll–Paque (GE Healthcare, Chicago, Ill). Dead cells were excluded with the LIVE/DEAD Fixable Aqua Dead Cell Stain (Invitrogen, Carlsbad, CA). After blocking with Fc receptor blocking solution (Biolegend, San Diego, CA), we incubated cells with 0.5 µg biotinylated human Dsg3 protein (CUSABIO, Wuhan, China) at 4°C for 1 h and made Dsg3 protein tetramers using streptavidin-BV421 and PE (Invitrogen). Then, the cells were stained with fluorescent-conjugated monoclonal antibody against CD3-BV510 (UCTH1), CD10-AF700 (HI10a), CD11c-BV605 (B-ly6), CD14-BV510 (MφP9), CD19-APC-Cy7 (SJ25C1), CD20-PerCP-Cy5.5 (2H7), CD27-BV711 (M-T271), CD38-PE-Cy7 (HIT2), IgD-BV786 (IA6-2) (all from BD Biosciences, Franklin Lakes, NJ), CD24-PE-CF594 (ML5) (Biolegend), and FcRL5-APC (509F6) (Invitrogen). Cells were detected with a BD LSRFortessa™ X-20 Cell Analyzer (BD Biosciences). For flow cytometric analysis, we have employed an established flow cytometry gating strategy as previously described (13 (link)) to detect antigen-specific B cells and atypical memory B cells. The data were analyzed by the FlowJo software package (BD Biosciences), and plots were generated using GraphPad Prism 9 (GraphPad Software Inc., San Diego, CA, USA).

Song A., Jang J., Lee A., Min S.Y., Lee S.G., Kim S.C., Shin J, & Kim J.H. (2022). Clinical impact and a prognostic marker of early rituximab treatment after rituximab reimbursement in Korean pemphigus patients. Frontiers in Immunology, 13, 932909.

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Comprehensive Lung Immune Cell Profiling

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Single cell suspensions were prepared by digesting lungs for 30 min in a solution of Collagenase II (Thermo Fisher Scientific) and DNase I (Roche Applied Science). Total lung cell counts were measured using a hemocytometer. Cells were stained for viability using LIVE/DEAD^® Fixable Aqua Dye (Life Technologies, A10346) and incubated with anti-CD16/CD32 (BioLegend, 101302) to block Fc receptors. Cells were then stained using fluorochrome conjugated anti-bodies to the following antigens: from BD Biosciences (San Jose, CA), Siglec-F-PE-CF594 (562757); and from BioLegend (San Diego, CA), CD45-APC-Cy7 (103116), CD11b-PerCP (101230), CD11c-BV605 (117334), Ly6C-BV711 (128037), Ly6G-BV421 (127628), CD64-PE (139304), F4/80-APC (123116), CD3-PE (100206), CD4-BV605 (100548), CD8-BV711 (100759), CD19-BV421 (115549), B220-PE Dazzle 594 (103258), and γδTCR-PerCP_Cy5.5 (118118).
Sample data were acquired on a BD LSR II equipped with FACSDiva™ software. All data analysis was performed using Flowjo™ software (Tree Star Inc.). Lung cell counts were expressed as the percentage of CD45⁺ live single cells. Cell subsets of neutrophils, (CD11b⁺, Ly6G⁺, F4/80⁻, CD64⁻), Ly6C⁺ monocytes (CD11b⁺, Ly6G⁻, Ly6C⁺, SiglecF⁻), alveolar macrophages (CD64⁺, CD11c⁺, SiglecF⁺), and γδTCR T cells (CD3⁺, B220⁻, γδTCR⁺) were defined using the above antibody combinations [14 (link)].

McGrath-Morrow S.A., Ndeh R., Collaco J.M., Poupore A.K., Dikeman D., Zhong Q., Singer B.D., D’Alessio F, & Scott A. (2017). The innate immune response to lower respiratory tract E. Coli infection and the role of the CCL2-CCR2 axis in neonatal mice. Cytokine, 97, 108-116.

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Multicolor Flow Cytometry Analysis of BALF

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Cells in BALF were stained with fluorescently labeled antibodies: CD45-Alexa Fluor 700 (30-F11), CD11b-PE-Cy7 (M1/70), Ly6G-PerCP-Cy5.5 (1A8), CD11c-BV605 (N418), CD45-Alexa Fluor 700 (30-F11), BV510-CD103 (2E7), SiglecF-Alexa Fluor 647 (E50-2440; BD Biosciences), MHCII-APC-Cy7 (M5/114.15.2), Ly6C-PE-Texas red (AL-2; BD Biosciences), and DAPI to assess cell viability. Uniform dyed microspheres, Dragon Green (Bangs Laboratories, Inc.), were used for cell counting. Antibodies were purchased from BioLegend. Data were collected by flow cytometry using an LSRFortessa X-20 cytometer (Becton Dickinson), and results were analyzed using FlowJo v.10.

Akoolo L., Pires S., Kim J, & Parker D. (2022). The Capsule of Acinetobacter baumannii Protects against the Innate Immune Response. Journal of Innate Immunity, 14(5), 543-554.

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