Levels of Th cells were determined as the percentage of CD4+ T cells that produce these cytokines following in vitro stimulation with PMA & Ionomycin. PBMCs, isolated as described above, were resuspended to 1×106 cells/mL in RPMI 1640 medium plus 10% FBS (R10). Cells were then incubated for 5h at 37°C in a medium with 1 × Cell Stimulation Cocktail (containing 81 nM PMA and 1.34 nM ionomycin plus protein transport inhibitors, Ebioscience, San Diego, CA, USA). Following incubation, the cells were washed and surface-stained with CD3-BV786, CD4-APC-Fire750, CD8-BV510 for 30 minutes in the dark at room temperature, followed by fixation and permeabilization. After permeabilization, cells were stained with IFN-γ-AF700 (clone 4S.B3), IL-22-FITC (clone 22URTI), IL-9-PE (clone MH9D1), IL-27-APC (clone ebic6), IL-10-PE-CY7 (clone JES3-9D7, Ebioscience), TNF-α-BV711 (clone MAb11), IL-17-BV421 (clone BL168), IL-2-BV650 (clone MQ1-17H12, BioLegend), or GM-CSF-PE-CF594 (clone BVD2-21C11, BD Biosciences) antibodies for 30 minutes in the dark at room temperature. Following staining, cells were washed and acquired on an LSRFortessa.
Il 10 pe cy7 clone jes3 9d7
Manufactured by Thermo Fisher Scientific
The IL-10-PE-CY7 is a fluorescent-labeled monoclonal antibody that binds to the human interleukin-10 (IL-10) protein. It is designed for use in flow cytometry applications to identify and quantify IL-10-expressing cells.
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Lab products found in correlation
2 protocols using il 10 pe cy7 clone jes3 9d7
1
Multiparameter T Cell Phenotyping
2
Flow Cytometric Analysis of IL-10 Expression in Stimulated PBMCs
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PBMCs, isolated as described above, were resuspended to 2×106 cells/mL in RPMI 1640 medium plus 10% FBS (R10). Cells were then incubated for 5h at 37°C in a medium with 1 × Cell Stimulation Cocktail (containing 81 nM PMA and 1.34 nM ionomycin plus protein transport inhibitors, Ebioscience). Following incubation, the cells were washed and surface-stained with CD3-BV786 (clone SK7), CD8-BUV395 (clone RPA-T8, BD Biosciences), CD4-APC-fire750 (clone SK3, BioLegend) for 30 minutes in the dark at room temperature, followed by fixation and permeabilization. After permeabilization, cells were stained with IL-10-PE-cy7 (clone JES3-9D7, Ebioscience) antibodies for 50 minutes in the dark at room temperature. A fixable viability dye eFluor 506 (Ebioscience) was used to assess cell viability. Following staining, cells were washed and acquired on an LSRFortessa.
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