NKp44+ ILCs from MLN of SIV-naïve rhesus macaques were stimulated overnight with either rhesus macaque IL-12 (50 ng/ml), human IL-1β (50 ng/ml), human IL-2 (1000 IU/ml) human IL-15 (50 ng/ml), or human IL-23 (50 ng/ml) (all from R&D Systems). After culture, cells were analyzed for intracellular expression of caspase-3 or RORγt (clone AFKJS-9, eBioscience).
Human il 23
Manufactured by R&D Systems
Human IL-23 is a recombinant human cytokine. It is a heterodimeric protein composed of two disulfide-linked subunits, p19 and p40. Human IL-23 is a member of the IL-12 family of cytokines.
Automatically generated - may contain errors
Lab products found in correlation
Human il 1β, by R&D Systems (2 mentions)
Human il 6, by R&D Systems (2 mentions)
Human tgf β, by R&D Systems (2 mentions)
Human il 2, by R&D Systems (1 mentions)
Human il 15, by R&D Systems (1 mentions)
Recombinant human il 12, by R&D Systems (1 mentions)
Purified anti human cd3 okt3, by Thermo Fisher Scientific (1 mentions)
Purified anti human cd28 cd28.2, by Thermo Fisher Scientific (1 mentions)
Mp4 25d2, by BioXCell (1 mentions)
Mouse il 6, by Thermo Fisher Scientific (1 mentions)
3 protocols using human il 23
1
Stimulation of NKp44+ ILCs from Rhesus Macaques
2
Detailed Immunology Protocol with Antibodies
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Purified anti-human CD3 (OKT3) and purified anti-human CD28 (CD28.2) were from eBioscience. Anti-human Bcl6 (EP529Y) was from abcam (Cambridge, MA, USA). Anti-CBP (D6C5) and anti-P300 (E6D1T) were from Cell Signal Technology (USA). Anti-β-actin (AB21800) and HRP-conjugated anti-rabbit and anti-mouse IgG (ABL3012 and ABL3032) were from Absci (Baltimore, MD, USA). Anti-β-hydroxybutyryllysine antibody (PTM-1204) was from PTMbio (China). Protein A/G-PLUS-agarose beads (37478) were from Cell Signaling Technology (Danvers, USA). Fluorochrome-conjugated anti-CD4 (RPA-T4), anti-human CD185 (RF8B2), anti-human CD279 (MIH4), and anti-human CD278 (DX29) were from BD Bioscience. Recombinant human IL-12, human TGFβ, human IL-23, and human IL-6 were from R&D Systems.
3
Th17 Cell Differentiation Modulation
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Purified naive CD4+ T cells (2 to 4 ×105 per well) were cultured under Th17 differentiation conditions in 96-well round-bottom plates. Cells collected from mice were cultured in medium containing 5 μg/mL anti-mouse CD3 mAb (17A2, BioXcell), 1 μg/mL anti-mouse CD28 mAb (37.51.1, eBioscience), 50 μM 2-mercaptoethanol (S6250, Sigma), 5 ng/mL human TGF-β (R&D), 20 ng/mL mouse IL-6 (Peprotech), 5 μg/mL anti-mouse IFN-γ mAb (XMG1.2, BioXcell), and 5 μg/mL anti-mouse IL-4 mAb (11B11, BioXcell) for 3 days. Cells collected from humans were cultured in medium containing 5 μg/mL anti-human CD3 mAb (OKT3, eBioscience), 1 μg/mL soluble anti-human CD28 mAb (CD28.2, eBioscience), 50 μM 2-mercaptoethanol (Sigma), 2.5 ng/mL human TGF-β (R&D), 20 ng/mL human IL-6 (R&D), 10 ng/mL human IL-23 (R&D), 10 ng/mL human IL-1β (R&D), 5 μg/mL anti-human IFN-γ mAb (285-IF-100, R&D), and 5 μg/mL anti-mouse IL-4 mAb (MP4-25D2, BioXcell) for 6–7 days.
To determine the role of MDSCs and polyamines in Th17 cell differentiation, MDSCs (at a 1:1 ratio to naive CD4+ T cells) were added in the presence or absence of ornithine (ORN, 100 nM, Sigma), difluoromethylornithine (DFMO, 20 nM, Tocris), putrescine (PUT, 2.5 μM, Sigma), spermidine (SPD, 2.5 μM, Sigma), or spermine (SPM, 2.5 μM, Sigma).
To determine the role of MDSCs and polyamines in Th17 cell differentiation, MDSCs (at a 1:1 ratio to naive CD4+ T cells) were added in the presence or absence of ornithine (ORN, 100 nM, Sigma), difluoromethylornithine (DFMO, 20 nM, Tocris), putrescine (PUT, 2.5 μM, Sigma), spermidine (SPD, 2.5 μM, Sigma), or spermine (SPM, 2.5 μM, Sigma).
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