PMA and Ionomycin were purchased from Sigma-Aldrich and used for non-specific stimulation and the positive controls. Mouse anti-human CD3-APC CY7 (clone SK7) and anti-IFNγ-APC were from BD Biosciences. Mouse anti-human CD8-PerCP (clone RPA-T8) was from BIOLEGEND. Mouse anti-human IL-2-allophycocyanin (clone 17-7029-71), anti-CD45RA-FITC (clone HI100), anti-CCR7-PE CY7 (clone 3D12), anti-TNFα-PE (clone MAB11) and anti-IL-17-PE (clone BL 168) were from eBioscience. Anti-human HLA-A2-PE (clone BB7.2) was from Abcam. CMV-EBV-Flu peptide pool was used for the study of antigen specific CD8 T cell responses as described previously (22 (link)). Peptides were synthesized by the GeneImmune (Suzhou, China), consisting of 23 HLA class I-restricted T cell epitopes from well-defined CMV, EBV and FLU viral proteins. The peptides were restricted by the most common HLA alleles including HLA-A1, A2, A3, A11, A24, A68, B7, B8, B27, B35, B44 (Supplementary Figure 4A ) and pooled into 12 groups as indicated in Supplementary Figure 4B . Function-graded mouse anti-human CD28 and anti-CD49d antibodies were from BD Biosciences.
Anti tnfα pe clone mab11
Manufactured by Thermo Fisher Scientific
Anti-TNFα-PE (clone MAB11) is a fluorescently labeled antibody that binds to the tumor necrosis factor alpha (TNFα) protein. It is used in flow cytometry and other applications to detect and measure the presence of TNFα in biological samples.
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Lab products found in correlation
Anti ifn γ apc, by BD (1 mentions)
Phorbol myristate acetate (pma), by Merck Group (1 mentions)
Ionomycin, by Merck Group (1 mentions)
Anti cd49d, by BD (1 mentions)
Cba human inflammatory cytokine kit, by BD (1 mentions)
Anti cd14 fitc, by Beckman Coulter (1 mentions)
Permeabilization buffer, by Thermo Fisher Scientific (1 mentions)
Brefeldin a, by Thermo Fisher Scientific (1 mentions)
2 protocols using anti tnfα pe clone mab11
1
Comprehensive Characterization of Antigen-Specific T Cells
2
Intracellular Cytokine Measurement in Macrophages
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Intracellular tumour necrosis factor (TNF)‐α production was measured in unstimulated and stimulated macrophages with LPS (1 ng/ml) or LTA (1 µg/ml) for 4 h in the presence of brefeldin A (eBioscience). After stimulation, cells were stained with anti‐CD14‐FITC (Beckman Coulter) for 10 min at 4°C and afterwards fixed with intracellular (IC) fixation buffer for 20 min. Cells were then permeabilized using permeabilization buffer (eBioscience), stained subsequently with anti‐TNF‐α‐PE (clone Mab11, eBioscience) for 20 min and analysed by flow cytometer.
In addition, TNF‐α and TGF‐β were measured via enzyme‐linked immunosorbent assay (ELISA) from the cell culture supernatants after 4 h of stimulation. Additionally, cytokine levels of interleukin (IL)‐1β, IL‐6, IL‐8, IL‐10, TNF‐α and IL‐12p70 were analysed using the human inflammatory cytokine CBA kit (BD Biosciences) after 24 h of stimulation.
In addition, TNF‐α and TGF‐β were measured via enzyme‐linked immunosorbent assay (ELISA) from the cell culture supernatants after 4 h of stimulation. Additionally, cytokine levels of interleukin (IL)‐1β, IL‐6, IL‐8, IL‐10, TNF‐α and IL‐12p70 were analysed using the human inflammatory cytokine CBA kit (BD Biosciences) after 24 h of stimulation.
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