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Siglec f

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Siglec F is a cell surface receptor that belongs to the Siglec family. It is primarily expressed on eosinophils and plays a role in the regulation of eosinophil function.

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Lab products found in correlation

Cd11b, by BioLegend (5 mentions) Facscanto 2, by BD (4 mentions) F4 80, by BioLegend (3 mentions) Cd103, by BioLegend (3 mentions) Lsrfortessa, by BD (2 mentions) Lsrii platform, by BD (2 mentions) Mhcii, by BioLegend (2 mentions) Alexa fluor 647 conjugated goat anti rabbit igg, by Thermo Fisher Scientific (1 mentions) Anti mouse cd16 cd32, by BioLegend (1 mentions) Cytofix cytoperm kit, by BD (1 mentions) Seven amino actinomycin d, by BioLegend (1 mentions) Rabbit anti collagen type 1, by Rockland Immunochemicals (1 mentions) Dihydroethidium dhe, by Thermo Fisher Scientific (1 mentions) Lysmcre cre mice, by Jackson ImmunoResearch (1 mentions) Cd200r, by BioLegend (1 mentions) Mitosox, by R&D Systems (1 mentions) Lsrfortessa flow cytometer, by BD (1 mentions) Collagenase d, by Roche (1 mentions) Flow cytometry, by BD (1 mentions) Rorγt, by BD (1 mentions)

Close spelling variants of this product

Siglec‐F (E50‐2440) (4 citations) Rat anti-mouse Siglec F-APC (4 citations) PE Siglec-F (3 citations) Siglec-F-PE/CF594 (2 citations) Siglec F APC-Cy7 (2 citations) PE-conjugated anti-Siglec-F (2 citations)

16 protocols using siglec f

1

Multiparametric Flow Cytometry Analysis

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Isolated cells were incubated with anti-mouse CD16/CD32 (BioLegend) to block non-specific Fc receptors. Then, the cell surface was stained with the corresponding mixture of fluorescently labelled monoclonal antibodies against B220, CD3, CD11b, CD45, F4/80, MHC II (IA/IE), Ly6C, Ly6G, Siglec F (BioLegend), NK-1.1 (Miltenyi Biotec, Auburn, CA, USA) and CCR2. The lineage cocktail consisted of antibodies targeting B220, CD3, Ly6G, NK1.1 and Siglec F. Seven-amino actinomycin D (BioLegend) was used to discriminate live and dead cells. Isolated cells were first gated on size, singularity and positive expression of EGFP and CD45. Next, lineage- and Seven-amino actinomycin D-positive cells were eliminated. To stain the intracellular antigens after surface labelling, cells were fixed and permeabilised with Cytofix/Cytoperm kit (BD Biosciences, San Diego, CA, USA) and sequentially incubated with rabbit anti-collagen type I (Rockland, Limerick, PA, USA) and Alexa Fluor 647-conjugated goat anti-rabbit IgG (Invitrogen). Data were acquired on LSRFortessa (BD Biosciences) and processed using FlowJo software (Tree Star, Ashland, OR, USA) with the appropriate isotype controls to determine the gating.
Hachiya K., Masuya M., Kuroda N., Yoneda M., Tsuboi J., Nagaharu K., Nishimura K., Shiotani T., Ohishi K., Tawara I, & Katayama N. (2021). Irbesartan, an angiotensin II type 1 receptor blocker, inhibits colitis-associated tumourigenesis by blocking the MCP-1/CCR2 pathway. Scientific Reports, 11, 19943.
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2

Macrophage ROS Metabolism Regulation

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All the mice used here have the C57BL/6 background. Atg7fl/fl mice were described previously (20 (link)). LysMcre/cre mice were purchased from Jackson Laboratories. All the experiments were performed as approved by IACUC. Antibodies against CD45, CD11b, F4/80, TLR4, TLR2, CD4, CD3, CD11c, Siglec F, CD200R, Ly6G and Ly6C were purchased from BioLegend. Dectin-1 and dectin-2 antibodies were from AbD Serotec. Antibodies against TREM2 and MARCO were purchased by R&D. MitoSOX and Dihydroethidium (DHE) were purchased from Life Technologies. An ROS inhibitor, N-acetyl-L-cyteine (NAC) was purchased from Sigma.
Kanayama M., He Y.W, & Shinohara M.L. (2015). Lung is protected from spontaneous inflammation by autophagy in myeloid cells. Journal of immunology (Baltimore, Md. : 1950), 194(11), 5465-5471.
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3

Isolation and Analysis of Lung and Spleen Immune Cells

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Mice were injected i.v. with 1 µg anti-CD8β 3 min before tissue harvest. Cells in the lung airways were recovered by lavage with PBS (5 × 1 ml), followed by plastic adherence (1 h at 37°C). Lung tissues were digested by collagenase D (Roche Diagnostics) for 30 min at 37°C, and enriched by centrifugation over a 40/80% Percoll gradient. Spleen cells were obtained by straining through nylon mesh and enriched by panning on anti-mouse IgG-coated flasks, followed by RBC lysis in buffered ammonium chloride. Cells were blocked first with mAbs to FcRγIII/II and then stained with APC-conjugated influenza NP366-374/Db tetramer. MHC class I tetramers were generated by the Molecular Biology Core Facility at the Trudeau Institute as described previously (Takamura et al., 2010 (link)). Tetramer-labeled cells were washed and stained with fluorescent-conjugated reagents purchased from BD Bioscience (CD45.1, CD90.1, Siglec F, and CD31), BioLegend (CD8α, CD8β, CD44, CD69, CD49a, CD103, CD11a, EpCAM, CCR3, CCR5, CCR6, and CCR9), TONBO Bioscience (CD90.2, CD11c, and F4/80), or R&D Systems (CXCR3, CXCR4, and CXCR6). Samples were run on LSRFortessa flow cytometers (BD Bioscience), and data were analyzed using FlowJo (Tree Star).
Takamura S., Kato S., Motozono C., Shimaoka T., Ueha S., Matsuo K., Miyauchi K., Masumoto T., Katsushima A., Nakayama T., Tomura M., Matsushima K., Kubo M, & Miyazawa M. (2019). Interstitial-resident memory CD8+ T cells sustain frontline epithelial memory in the lung. The Journal of Experimental Medicine, 216(12), 2736-2747.
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4

Colonic Immune Cell Profiling by Flow Cytometry

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The colonic single cells of lamina propria were suspended and stained with mouse monoclonal antibodies, including CD45 (BioLegend, #103132), CD11b (BioLegend, #101206), F4/80 (BioLegend, #123116), CD3 (BioLegend, #100305), B220 (BioLegend, #103223), Ly6C (BioLegend, #128026), CX3CR1 (BioLegend, #149006), SiglecF (BioLegend, #155505), Ly6G (BioLegend, #127613), CD45 (BD, #563891), CD3e (BD, #553061), CD4 (BD, #552051), CD8a (BD, #553030), RORγt (BD, #563081), T-Bet (BD, #563318), and Foxp3 (BD, #560414). The antibodies were incubated at 4 °C in the dark for 30 min. The cell subsets classification was detected by Flow cytometry (BD Celasta). Flow cytometric data were analyzed by using FlowJo software (BD).
Yang L., Shen W.W., Shao W., Zhao Q., Pang G.Z., Yang Y., Tao X.F., Zhang W.P., Mei Q, & Shen Y.X. (2023). MANF ameliorates DSS-induced mouse colitis via restricting Ly6ChiCX3CR1int macrophage transformation and suppressing CHOP-BATF2 signaling pathway. Acta Pharmacologica Sinica, 44(6), 1175-1190.
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5

Comprehensive Mouse Spleen and Bone Marrow Immunophenotyping

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Briefly, mouse spleens and BM were collected and dissociated through a 40μm strainer to produce a single cell suspension. Red blood cells were lysed using 0.86% ammonium chloride (Sigma). Cells were counted, washed in flow cytometry buffer (1% BSA (Sigma), 2mM EDTA (Invitrogen) in PBS. FcR were blocked with 2.4G2 (BioXCell) and Rat IgG (Invitrogen) while staining with McAb at 40 for 30 minutes. Intracellular staining was carried out using the eBioscience kit. Samples were acquired using a FACSCanto II or LSRFortessa flow cytometer (BD Biosciences) and analyzed with FlowJo software (Tree Star, Inc.). The following mAb against mouse antigens were used as FITC, PE, PerCP-Cy5.5, PE-Cy7, allophycocyanin (APC), APC-ef780, Pacific blue, AF450, AF700, PE Texas Red, or biotin conjugates: FcεRI, AA4.1, CD23, CD43, IgM, FcγRII/III, ckit, Sca-1, CD34, CD138, CD45R (B220; RA3-6B2), CD278 (ICOS; C398.4A), IgD (11–26) (eBioscience), CD4 (RM4–5), CD8a (53–6.7), CD95 (Fas; Jo2), CXCR5 (2G8) (BD Biosciences), CD3 (17A2), CD19 (6D5), Foxp3, CTLA-4, Siglec F, CD11b, F4/80, TCRβ, Gr-1, FcεRI, and CD279 (PD-1; J43) (Biolegend). Biotinylated antibodies were detected using PerCP-Cy5.5– (BD Biosciences) or APC-conjugated streptavidin (eBioscience). FITC or biotin-conjugated PNA was obtained from Vector Laboratories. Plots shown are on a Logicle scale.
Zaretsky A.G., Konradt C., Dépis F., Wing J.B., Goenka R., Atria D.G., Silver J.S., Cho S., Wolf A.I., Quinn W.J., Engiles J.B., Brown D.C., Beiting D., Erikson J., Allman D., Cancro M.P., Sakaguchi S., Lu L., Benoist C.O, & Hunter C.A. (2017). T Regulatory Cells Support Plasma Cell Populations in the Bone Marrow. Cell reports, 18(8), 1906-1916.
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6

Flow Cytometric Analysis of Lung Leukocytes

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The cells yielded from the BAL and lung were stained using fluorescent labeled antibodies at 4°C in the dark for 20 minutes, washed with PBS, and then examined on the BD Biosciences (San Jose, CA) LSR II platform in the BRI flow cytometry core.
Leukocytes were stained for CD11b, CD11c, CD45.2, CD64, CD103, Ly6C, Ly6G, MHC-II, SiglecF (all Biolegend), CD253 (TRAIL), and Viability (eBioscience). The eBioscience (ThermoFisher Scientific, Foster City, CA)) Annexin V Apoptosis Detection Kit was used in accordance with the manufacturer’s instructions to determine which of the cells that absorbed the Viability dye had undergone a programmed cell death process. (See Supplemental Fig.1 for the complete staining and pictorial gating scheme.)
Kellar G.G., Reeves S.R., Barrow K.A., Debley J.S., Wight T.N, & Ziegler S.F. (2020). Juvenile, but not adult, mice display increased myeloid recruitment and extracellular matrix remodeling during respiratory syncytial virus infection. Journal of immunology (Baltimore, Md. : 1950), 205(11), 3050-3057.
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7

Isolation and Characterization of Alveolar Macrophages

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BALF was collected as described (Li et al, 2013 (link)) and centrifuged at 400g for 10 min to separate the supernatant from cells. The numbers of total cells, neutrophils, and macrophages from BALF were visualized using the Wright–Giemsa staining (Sigma-Aldrich) and counted in a double-blind manner. Isolation of alveolar macrophages from BALF was performed as described in detail earlier (Busch et al, 2019 (link)), and the purity of isolated macrophages was characterized by dual staining with PE-conjugated sialic acid–binding immunoglobulin-like lectin F (Siglec-F) and Alexa Fluor 488–conjugated CD11c (BioLegend) followed by flow cytometry.
Gao P., Wu B., Ding Y., Yin B, & Gu H. (2022). circEXOC5 promotes acute lung injury through the PTBP1/Skp2/Runx2 axis to activate autophagy. Life Science Alliance, 6(1), e202201468.
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8

Comprehensive Immune Cell Profiling

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Single-cell suspensions were stained on ice with the following eBioscience antibodies: B220 (RA3-6B2), Siglec-F (1RNM44N), CD103 (2E7), CD11b (MI-70), CD11c (N418), CD8α (53-6.7), CD69 (H1-2F3), CD4 (RM4-5), CD3ε (145-2C11), γδ T cell receptor (γδTCR) (GL3), NK1.1 (PK136), CD44 (IM7), Ly6G (1A8), Ly6C (HK1.4), MHCII (M5), and XCR1 (ZET; BioLegend). Intracellular IgG2c was detected with polyclonal goat antiserum (Southern Biotech) using Cytofix/Cytoperm (Becton Dickinson). Analysis was performed on LSR II or FACSCanto II (Becton Dickinson) flow cytometers. Data were analyzed with FlowJo (TreeStar).
Vogelzang A., Lozza L., Reece S.T., Perdomo C., Zedler U., Hahnke K., Oberbeck-Mueller D., Dorhoi A, & Kaufmann S.H. (2016). Neonatal Fc Receptor Regulation of Lung Immunoglobulin and CD103+ Dendritic Cells Confers Transient Susceptibility to Tuberculosis. Infection and Immunity, 84(10), 2914-2921.
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9

Multiparameter Flow Cytometry of Immune Cells

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Flow cytometric analysis was performed on a Canto II flow cytometer (BD Biosciences) using FlowJo software v10.7.1 (Tree Star Inc., Ashland, OR, USA). Dead cells were excluded using a live/dead fixable dead cell stain (Thermo Fisher Scientific, Waltham, MA, USA). The following mouse-specific antibodies were used for staining: CD3 (145-2C11, BioLegend, San Diego, CA, USA), CD4 (RM4-5, BioLegend), Foxp3 (FJK-16s, Invitrogen), T-bet (4B10, Invitrogen, Eugene, OR, USA), RORγt (B2D, Invitrogen), GATA3 (16E10A23, BioLegend), Siglec F (S17007L, Biolegend), Ly6G (1A8, Biolegend), CD11b (M1/70, BioLegend), B220 (RA3-6B2, BioLegend), IgA (C10-3, Biolegend), and IgM (RMM-1, Biolegend).
Gu B.H., Rim C.Y., Lee S., Kim T.Y., Joo S.S., Lee S.J., Park H.K, & Kim M. (2022). Alteration of Gut Immunity and Microbiome in Mixed Granulocytic Asthma. Biomedicines, 10(11), 2946.
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10

Isolating Murine Lung Cell Populations

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Mouse lung tissues were ground into single cell suspensions using a Miltenyi automatic tissue grinder (Miltenyi, Germany). The flow cytometry antibodies used were CD11c (117309, Biolegend, USA) and Siglec F (155509, Biolegend, USA). Cell surface staining was performed at 4°C for 30–40 min in the dark. The purity of sorted populations was >99%, unless otherwise indicated. Data were obtained on a FACSCanto II clinical flow cytometry system (BD Biosciences) and analyzed using FlowJo software (FlowJo LLC, Ashland, Oregon).
Yu L., Wang L., Hu G., Ren L., Qiu C., Li S., Zhou X., Chen S, & Chen R. (2022). Reprogramming alternative macrophage polarization by GATM-mediated endogenous creatine synthesis: A potential target for HDM-induced asthma treatment. Frontiers in Immunology, 13, 937331.
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