All FACS antibodies were purchased from eBioscience. For immunohistochemistry FITC-conjugated anti-B220 mAb and biotinylated anti-IgD were purchased from eBiosciences streptavidin-conjugated alexafluor594 was purchased from Molecular Probes. Surface stainings were performed as previously described45 (link). For intracellular staining, cells were stained then fixed/permeabilized according to the eBioscience protocol. Permeabilized cells were incubated in Permwash™ with anti-IL10, anti-TNF, or appropriate isotype control for 30 mins at 4 °C (with blocking anti-CD16/32). Intracellular staining for FOXP3 was performed with eBioscience FOXP3 kit according to manufacturer’s instructions. For the detection of intracellular cytokines PMA (50 ng/ml), ionomycin (1 μM) and brefeldin A were added for 5 hrs before staining. Cells were acquired using LSRII or Fortessa flow cytometers (BD Biosciences). Flow Jo software was used for analysis.
Facs antibodies
Manufactured by Thermo Fisher Scientific
Sourced in United States
FACS antibodies are used in flow cytometry applications to label and identify specific cell types or cellular markers. They are conjugated with fluorescent dyes that allow the detection and quantification of target cells or molecules within a sample.
Automatically generated - may contain errors
Lab products found in correlation
Fortessa flow cytometer, by BD (1 mentions)
Fortessa, by BD (1 mentions)
Ki 67 rm 9106 s1, by Thermo Fisher Scientific (1 mentions)
Anti cd3, by Jackson ImmunoResearch (1 mentions)
Mouse neutrophil isolation kit, by Miltenyi Biotec (1 mentions)
Rs504393, by Merck Group (1 mentions)
Ly6g antibody, by BioLegend (1 mentions)
Cxcl5, by R&D Systems (1 mentions)
Sb431542, by Merck Group (1 mentions)
Cxcl1, by R&D Systems (1 mentions)
Bouin s solution, by Merck Group (1 mentions)
5 protocols using facs antibodies
1
Multiparametric Flow Cytometry Analysis
2
Intracellular Cytokine Staining Protocol
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All FACS antibodies were purchased from eBioscience. Surface staining was performed as previously described39 (link). For intracellular staining, cells were stained then fixed/permebilized according to the eBioscience protocol. Permeabilized cells were incubated in PermwashTM with anti-IL10, anti-TNF-α or appropriate isotype control for 30 mins at 4 °C (with blocking anti-CD16/32). For the detection of intracellular cytokines, PMA (50ng/ml), ionomycin (1 μM) and brefeldin A were added for 5 hrs before staining. Cells were acquired using LSRII or Fortessa flow cytometers (BD Biosciences). FlowJo software was used for analysis.
3
Multiparameter Cytokine Profiling of Lung Leukocytes
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For intracellular cytokine staining, isolated lung leukocytes were stimulated with soluble αCD3 and αCD28 (both at 1 μg/ml) in the presence of brefeldin A and monensin for 5 hours. Lung cells were stained with various combinations of the following mAb clones: αTNF MP6-XT22, αCD8 53.67, αIL-17A TC11-18H10.1, αCD4 RM4-5, αCD45.2 104, αIL-5 TRFK5, αCD44 IM7, αIFNγ XMG1.2, αIL13 eBio13A, αFoxp3 FJK-16S, αLy6G 1A8, αCD11b M1/70, αCD11c N418, αCD103 2E7, αCD68 FA11, αLy6C HK1.4, αSiglec F E50-2440, αT-bet 4B10, αPD-1 29F.1A12, αRORγt B2D, and αGATA-3 TWAJ. All FACS antibodies were purchased from eBioscience (San Diego, CA) and Biolegend (San Diego, CA). Analysis of CD4 T cell cytokine polyfunctionality was performed using SPICE version 5.3, downloaded from http://exon.niaid.nih.gov [72 (link)].
4
Antibody-Based Tumor and Lineage Analysis
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Antibodies used for tumor analysis include F4/80 (ab6640, Abcam, Cambridge, MA, USA) [99 (link)], EphA2 (C-20, SC-924, Santa Cruz Biotech, Dallas, TX, USA), Ki-67 (RM-9106-S1, Thermo Fisher Scientific, Waltham, MA, USA), SREBP1 (H-160), SREBP2 (H-164), FASN (H-300), and CD31 (CM303A, Biocare Medical, Concord, CA, USA). Antibodies used for lineage analysis were: anti-B220 (RA3-6B2), anti-CD19 (eBio1D3), anti-IgM µ-chain (Jackson Labs), anti-CD43 (S7) for B cells, anti-CD3 (145-2C11), anti-CD4 (GK1.5), anti-CD8a (53-6.7) for T cells, anti-CD41 (MWReg30), anti-CD71 (C2), anti-Ter119 (TER-119), and anti-F4/80 (BM8), anti-Mac1 (M1/70), and anti-Gr1 (RB6-8C5) for myeloid cells. Antibodies used for HSC and progenitor analysis were: Lineage (biotin-conjugated anti-Gr-1 (RB6-8C5), -Mac1 (M1/70), -B220 (RA3-6B2), -CD19 (eBio1D3), -Ter110 (TER-119), -CD5 (53-7.3), -CD4 (GK1.5), -CD8 (53-6.7)), APC-Cy7-c-Kit (2B8), PerCP-cy5.5-Sca1 (E13-161.7 or D7, 1:1000 dilution), FITC-CD48 (HM48-1), PE-Cy7-CD150 (TC15-12F12.2), APC-CD34 (RAM34) and PE–Flk2 (A2F10.1). All antibodies were used at 1:200 dilution unless otherwise noted. FACS antibodies were purchased from eBioscience (San Diego, CA, USA), BD Biosciences (San Jose, CA, USA) or BioLegend (San Diego, CA, USA).
5
Inflammatory Mediator Detection
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Recombinant mouse TNFα, ELISA kit for testing mouse CCL5, CXCL1 and CXCL5 were purchased from R&D Systems (Minneapolis, MN, USA). FACS antibodies were from eBiosciences (La Jolla, CA, USA). Bouin's solution, RS504393 and SB431542 were from Sigma-Aldrich. Mouse neutrophil isolation kit was purchased from Miltenyi Biotec (Bergisch Gladbach, Germany). Ly6G antibody and isotype antibody were purchased from Biolegend (San Diego, CA, USA). PCR Array Kit for Mouse Chemokines & Receptors was purchased from SABiosciences (Valencia, CA, USA).
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