All cells were labeled with live/dead dye near infra red (Life Technologies) for dead cell exclusion and treated with Fc Block (Miltenyi, San Diego, CA, USA) prior to staining with fluorescently labeled antibodies. Anti-human antibodies used for DC staining were anti-CD83 PE (Biolegend, San Diego, CA, USA), anti-CD86-PE-Cy7 (Biolegend), anti-HLA-DR V450 (BD), and anti-CD11c APC (BD). Antibodies used in the T cell characterization were anti-LAG-3 FITC (Novus, Littleton, CO, USA), anti-PD-1 PerCP-Cy5.5, anti-CD3 Alexa700, anti-CD4 Brilliant Violet 650, anti-CD8 Brilliant Violet 570, anti-IFN-γ PE, anti-IL-5, anti-CD62L PE, and anti-CD45RA Alexa700 (all, Biolegend), anti-CD25 PE, and anti-IFN-γ PE-Cy7, anti-IL-4 PE-Cy7 (all BD), and anti-TIM-3 APC (R&D Systems). Surface staining and intracellular staining was performed using Cytofix/Cytoperm™ Plus kit (BD) according to the manufacturer’s instructions. Data acquisition was performed using the LSRFortessa flow cytometer (BD). Data analysis was performed with FlowJo version 9 (Tree Star, Inc., Ashland, OR, USA).
Anti tim 3 apc
Manufactured by R&D Systems
Sourced in United States, Germany
Anti-TIM-3 APC is a laboratory reagent used in flow cytometry analysis. It binds to the TIM-3 (T-cell immunoglobulin and mucin-domain containing-3) protein expressed on the surface of certain immune cells. This reagent can be used to identify and quantify TIM-3 positive cells in a sample.
Automatically generated - may contain errors
Lab products found in correlation
Lsrfortessa flow cytometer, by BD (1 mentions)
Anti cd25 pe, by BD (1 mentions)
Anti cd11c apc, by BD (1 mentions)
Anti ifn γ pe, by BioLegend (1 mentions)
Anti cd86 pe cy7, by BioLegend (1 mentions)
Pe anti cd62l, by BioLegend (1 mentions)
Fc block, by Miltenyi Biotec (1 mentions)
Cytofix cytoperm plus kit, by BD (1 mentions)
Anti pd 1 percpcy 5, by BioLegend (1 mentions)
Anti cd83 pe, by BioLegend (1 mentions)
Anti hla dr v450, by BD (1 mentions)
Anti cd3 alexa700, by BioLegend (1 mentions)
Anti cd4 brilliant violet 650, by BioLegend (1 mentions)
Anti il 5, by BioLegend (1 mentions)
Anti il 4 pe cy7, by BD (1 mentions)
Anti ifn γ pe cy7, by BD (1 mentions)
Ec800 flow cytometer, by Sony (1 mentions)
Rat igg2a, by R&D Systems (1 mentions)
Mouse igg2b, by R&D Systems (1 mentions)
Mouse igg1κ, by BD (1 mentions)
4 protocols using anti tim 3 apc
1
Multiparametric Flow Cytometry Analysis
2
Evaluating Cell Surface Antigen Expression
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Cell surface antigen expressions were analyzed using flowcytometry. The cells were incubated with anti-HER2-PE (clone; #191924, isotype; mouse IgG2b, R&D Systems, MN), anti-NKG2D-APC (clone; 1D11, isotype; mouse IgG1κ, BD Biosciences, Heidelberg, Germany), anti-Tim3-APC (clone; #344823, isotype; rat IgG2a, R&D systems) for 30 min at 4 °C. The cells were then washed and analyzed using an EC800 flowcytometer (SONY, Tokyo, Japan).
3
Comprehensive Immune Cell Profiling
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Venous blood samples (1 mL) were collected from individual patients and healthy individuals immediately after visiting the hospital. To determine the frequency of T cells (CD3/CD4/CD8), natural killer (NK) cells, and monocytes (CD14), blood samples were stained with the following antibodies: Pcy5-conjugated anti-CD3, FITC-conjugated anti-CD4, P-phycoerythrin-conjugated anti-CD8 (BD Biosciences, CA, USA), and PE-conjugated anti-CD16/CD56 (Beckman Coulter, CA, USA).
The frequency of different types of immunocompetent cells was characterized by flow cytometry as previously described (PMID: 8404602). Briefly, at least 1 × 105 cells were analyzed using BD FACS CantoTM II flow cytometer (BD Bioscience, CA, USA). The staining of T cells or monocytes was performed with a single antibody or a combination of the following antibodies: anti-Tim-3-APC (R&D Systems), anti-PD-1-PE (eBiosciences), anti-PD-L1-BrilliantViolet421 (Biolegend), and anti-PD-L2-APC (Biolegend). The isotype-matched immunoglobulins were used as controls. The tests were performed at least in triplicate.
The frequency of different types of immunocompetent cells was characterized by flow cytometry as previously described (PMID: 8404602). Briefly, at least 1 × 105 cells were analyzed using BD FACS CantoTM II flow cytometer (BD Bioscience, CA, USA). The staining of T cells or monocytes was performed with a single antibody or a combination of the following antibodies: anti-Tim-3-APC (R&D Systems), anti-PD-1-PE (eBiosciences), anti-PD-L1-BrilliantViolet421 (Biolegend), and anti-PD-L2-APC (Biolegend). The isotype-matched immunoglobulins were used as controls. The tests were performed at least in triplicate.
4
Multiparametric Flow Cytometry of Immune Cells
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Zombie UV™ Fixable Viability Kit, anti-CD45 BV570, anti-CD3 BV605, anti-CD4 BV421, anti-CD8 BV650, anti-CD4 APC-Fire750, anti-CD73 PE/Dazzle 594, anti-CD56 PE, anti-CD14 PerCP/Cy5.5, anti-CD16 Alex Fluor 700, anti-CD19 APC-Fire750, anti-HLA-DR PE-Cy5, anti-PD-1 BV421, anti-CD16 PE, anti-IFNγ A700, anti-IL-13 APC, anti-IL-5 APC, anti-TNFα Alexa Fluor 488, and anti-IL-21 PE were purchased from BioLegend (London, UK). Anti-CD25 BB515, anti-CD4 BUV496, anti-CD39 BV711, anti-Perforin Alexa Fluor 647, anti-Granzyme B Alexa Fluor 700, anti-Ki-67 Alexa Fluor 700, anti-CD45RA PE, anti-IL-2 PerCP/Cy5.5, and Fixation/Permeabilization Solution Kit were purchased from Becton Dickinson. Anti-FOXP3 APC, anti-CD56 APC-eFluor780, and eBioscience™ (San Diego, CA, USA) Foxp3/Transcription Factor Staining Buffer Set were purchased from eBioscience TermoFisher scientific. Anti-TIM-3 APC was purchased from R&D systems. Anti-CD137 PE and FcR blocking reagent were purchased from Miltenyi (Bergisch Gladbach, Germany).
Peptides for the PSA, PSMA, PSCA, and PAP pools (Supplementary Table S1 ) were synthetized and lyophilized by the Peptide and Tetramer Core Facility of the Department of Oncology at UNIL-CHUV (Lausanne, Switzerland), and 5T4 pools (Supplementary Tables S2 and S3 ) were synthetized and lyophilized by JPT Peptide Technologies.
Peptides for the PSA, PSMA, PSCA, and PAP pools (
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