T cells were re-stimulated with phorbol myristate acetate (Sigma, 50 ng/mL) and ionomycin (Sigma, 500 ng/mL). 90 minutes later, cells were treated with Brefeldin A to block cytokine secretion. 3 h later, cells were stained for surface markers and simultaneously labelled with Live/Dead Blue Viability Dye (Thermo Fisher) for 20 minutes at 4°C. Cells were washed twice and fixed overnight using a FoxP3 Fixation/Permeabilization kit (Thermo Fisher). The following day, cells were washed and stained for intracellular cytokines at room temperature for 1 h. They were then washed 3 times and analysed using an LSR Fortessa machine (Beckman Dickinson). Analysis of mean fluorescence intensity was performed using FlowJo v10.0. All experiments were performed at least two independent times. Antibodies used (at 1:100 unless otherwise noted) were TNF-PE (BioLegend, MP6-XT22, 506306), PD-1-PECy7 (BioLegend, RMP1–30, 109110) IFN-γ-FITC (BioLegend, XMG1.2, 505806), CD4-BV711 (BioLegend, RM4–5, 100550), CD8α-BV786 (BioLegend, 53–6.7, 100750), Tox-PE (Miltenyi, REA473, 130–120-716, 1:50), Tcf7-Alexa647 (Cell Signaling, C63D9, 6709), PD-L1-APC (BioLegend, 10F.9G2, 124312, 1:5000), LAG-3-PerCP-Cy5.5 (BioLegend, C9B7W, 125211), TIM-3-PE-Dazzle594 (BioLegend, B8.2C12, 134013).
Tnf pe
Manufactured by BioLegend
TNF-PE is a lab equipment product used for the detection and quantification of tumor necrosis factor (TNF) in samples. It utilizes a fluorescent dye, phycoerythrin (PE), to label anti-TNF antibodies, enabling their visualization and measurement.
Automatically generated - may contain errors
Lab products found in correlation
Pd1 pe cy7, by BioLegend (2 mentions)
Foxp3 fixation permeabilization kit, by Thermo Fisher Scientific (2 mentions)
Pd l1 apc, by BioLegend (2 mentions)
Live dead blue viability dye, by Thermo Fisher Scientific (2 mentions)
Cd8α bv786, by BioLegend (2 mentions)
Tox pe, by Miltenyi Biotec (2 mentions)
Tcf7 alexa647, by Cell Signaling Technology (2 mentions)
Lag 3 percp cy5, by BioLegend (2 mentions)
Tim 3 pe dazzle594, by BioLegend (2 mentions)
Ifn γ fitc, by BioLegend (2 mentions)
Cd4 bv711, by BioLegend (2 mentions)
Ionomycin, by Merck Group (2 mentions)
Igg1 pe, by BioLegend (1 mentions)
3 protocols using tnf pe
1
Multiparametric T cell analysis
2
Quantifying Monocyte-Derived TNF Production
Check if the same lab product or an alternative is used in the 5 most similar protocols
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4 × 106 Ficoll‐isolated PBMCs were seeded in a 24‐well plate in 2 ml RPMI‐1640 (+ 10% FCS, 2 mM l ‐glutamine, 1 mM Sodium‐Pyruvate, 50 μM β‐Mercaptoethanol) for 4 h in presence of 10 µg/ml Brefeldin A (B7651, SA). Cell surface was stained using CD11b‐PB antibody (dilution 1:200; 301315, BioLegend) for 20 min at 4°C in the dark. Intracellular TNF was stained using TNF‐PE (dilution 1:20; 502909, BioLegend) or IgG1‐PE (dilution 1:20, 400140, BioLegend) as isotype control for 30 min at 4°C in the dark.
3
Multiparametric T cell analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
T cells were re-stimulated with phorbol myristate acetate (Sigma, 50 ng/mL) and ionomycin (Sigma, 500 ng/mL). 90 minutes later, cells were treated with Brefeldin A to block cytokine secretion. 3 h later, cells were stained for surface markers and simultaneously labelled with Live/Dead Blue Viability Dye (Thermo Fisher) for 20 minutes at 4°C. Cells were washed twice and fixed overnight using a FoxP3 Fixation/Permeabilization kit (Thermo Fisher). The following day, cells were washed and stained for intracellular cytokines at room temperature for 1 h. They were then washed 3 times and analysed using an LSR Fortessa machine (Beckman Dickinson). Analysis of mean fluorescence intensity was performed using FlowJo v10.0. All experiments were performed at least two independent times. Antibodies used (at 1:100 unless otherwise noted) were TNF-PE (BioLegend, MP6-XT22, 506306), PD-1-PECy7 (BioLegend, RMP1–30, 109110) IFN-γ-FITC (BioLegend, XMG1.2, 505806), CD4-BV711 (BioLegend, RM4–5, 100550), CD8α-BV786 (BioLegend, 53–6.7, 100750), Tox-PE (Miltenyi, REA473, 130–120-716, 1:50), Tcf7-Alexa647 (Cell Signaling, C63D9, 6709), PD-L1-APC (BioLegend, 10F.9G2, 124312, 1:5000), LAG-3-PerCP-Cy5.5 (BioLegend, C9B7W, 125211), TIM-3-PE-Dazzle594 (BioLegend, B8.2C12, 134013).
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