The antibodies we used in this study were anti-mouse CD4, CD19, CD45, B220, CD3, F4/80, CD11c, CD11b (BD Bioscience), IL-10, IL-17, IFNγ, IL-27p28 and Foxp3 (eBioscience), eBi3 (R&D systems) and LIVE/DEAD Cell Viability Kits (Invitrogen). MLN or colonic LP cells were collected and incubated for 15 minutes at 4°C with anti-CD16/CD32 (BD Bioscience) and then for 20 minutes at 4°C with antibodies for cell surface and LIVE/DEAD Cell Viability Kits to evaluate the cell phenotype and CD4+ T cell/B cell reconstitution. Enumeration of cells expressing IL-10, IL-17, IFNγ, IL-27p28, Ebi3 and Foxp3 was performed by intracellular staining with a Fixation and Permeabilization Solution Kit (BD Bioscience) according to the manufacturer's instructions. For intracellular cytokine staining, 100ng/ml PMA, 1µg/ml Ionomycin (Sigma-Aldrich) and Golgi stop (BD Biosciences) were added into the medium during the last 4 hours of the culture period. Cells were washed and then analyzed on a CyAn flow cytometer (Beckman Coulter, Brea, CA, USA). Proper isotype antibodies were used as a control and gated live CD45+ cells were analyzed with Summit 5.2 software (Beckman Coulter)
Il 27p28
Manufactured by BD
IL-27p28 is a recombinant protein that represents the p28 subunit of the heterodimeric cytokine IL-27. IL-27 is a member of the IL-12 family of cytokines and plays a role in the regulation of immune responses.
Automatically generated - may contain errors
Lab products found in correlation
Cyan flow cytometer, by Beckman Coulter (2 mentions)
Anti mouse cd4, by BD (2 mentions)
Live dead cell viability kit, by Thermo Fisher Scientific (2 mentions)
Ionomycin, by Merck Group (2 mentions)
Golgistop, by BD (2 mentions)
Summit 5, by Beckman Coulter (2 mentions)
Fixation and permeabilization solution kit, by BD (2 mentions)
Live dead cell viability kits, by BD (2 mentions)
Anti cd16 cd32, by BD (2 mentions)
Cd11b, by BD (1 mentions)
F4 80, by BD (1 mentions)
2 protocols using il 27p28
1
Immune Cell Profiling by Flow Cytometry
2
Multicolor Flow Cytometry Analysis of Immune Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
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The antibodies we used in this study were anti-mouse CD4, CD19, CD45, B220, CD3, F4/80, CD11c, CD11b (BD Bioscience), IL10, IL17, IFNγ, IL27p28, and Foxp3 (eBioscience), eBi3 (R&D Systems, Minneapolis, MN), and live/dead cell viability kits (Invitrogen/Life Technologies, Carlsbad, CA). MLN, or colonic LP cells were collected and incubated for 15 minutes at 4°C with anti-CD16/CD32 (BD Bioscience) and then for 20 minutes at 4°C with antibodies for cell surface and live/dead cell viability kits to evaluate the cell phenotype and CD4+ T-cell/B-cell reconstitution.
Enumeration of cells expressing IL10, IL17, IFNγ, IL27p28, Ebi3, and Foxp3 was performed by intracellular staining with a fixation and permeabilization solution kit (BD Bioscience) according to the manufacturer’s instructions. For intracellular cytokine staining, 100 ng/mL phorbol myristate acetate, 1 μg/mL ionomycin (Sigma-Aldrich), and GolgiStop (BD Biosciences) were added into the medium during the last 4 hours of the culture period. The cells were washed and then analyzed on a CyAn flow cytometer (Beckman Coulter, Brea, CA). Proper isotype antibodies were used as a control, and gated live CD45+ cells were analyzed with Summit 5.2 software (Beckman Coulter).
Enumeration of cells expressing IL10, IL17, IFNγ, IL27p28, Ebi3, and Foxp3 was performed by intracellular staining with a fixation and permeabilization solution kit (BD Bioscience) according to the manufacturer’s instructions. For intracellular cytokine staining, 100 ng/mL phorbol myristate acetate, 1 μg/mL ionomycin (Sigma-Aldrich), and GolgiStop (BD Biosciences) were added into the medium during the last 4 hours of the culture period. The cells were washed and then analyzed on a CyAn flow cytometer (Beckman Coulter, Brea, CA). Proper isotype antibodies were used as a control, and gated live CD45+ cells were analyzed with Summit 5.2 software (Beckman Coulter).
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