CD4+ T cells were isolated using EasySep mouse CD4+ T cell isolation kit (Stemcell Technologies) with the addition of biotinylated anti-CD25 antibody (BioLegend). Cells were cultured in Iscove’s modified Dulbecco medium (IMDM, Sigma) supplemented with 2mM L-glutamine, 100 U/ml penicillin, 100 µg/ml streptomycin, 0.05mM β-mercaptoethanol and 5% fetal calf serum (biosera). CD4+ T cells were differentiated in 48 well plates coated with 1 µg/ml anti-CD3 (clone 145-2C11, eBioscience) and 10 µg/ml soluble anti-CD28 (clone 37.51, BioLegend) in the presence of 2ng/ml TGF-β1, 20ng/ml IL-6, 10ng/ml IL-1β (all R&D Systems) and 10μg anti-IFN-γ (BioXCell). In some cultures 6-formylindolo[3,2-b]carbazole (FICZ, EnzoLifeSciences) was added from the start of culture. IL-22 cytokine levels in culture supernatants were determined by ELISA (eBioscience).
Soluble anti cd28 clone 37
Manufactured by BioLegend
Soluble anti-CD28 (clone 37.51) is a monoclonal antibody that binds to the CD28 receptor on T cells. CD28 is a co-stimulatory receptor that plays a critical role in T cell activation and proliferation.
Automatically generated - may contain errors
Lab products found in correlation
Easysep mouse cd4 t cell isolation kit, by STEMCELL (2 mentions)
Anti ifn γ, by BioXCell (2 mentions)
Fetal calf serum (fcs), by Biosera (2 mentions)
Anti cd3 clone 145 2c11, by Thermo Fisher Scientific (2 mentions)
Interleukin 6 (il 6), by R&D Systems (2 mentions)
Biotinylated anti cd25 antibody, by BioLegend (2 mentions)
Il 1β, by R&D Systems (2 mentions)
Tgf β1, by R&D Systems (2 mentions)
Interleukin 7 (il 7), by Thermo Fisher Scientific (1 mentions)
Carboxyfluorescein succinimidyl ester (cfse), by BD (1 mentions)
3 protocols using soluble anti cd28 clone 37
1
CD4+ T Cell Differentiation Protocol
2
CD4+ T Cell Differentiation Protocol
3
T Cell Activation and Cytokine Modulation
Check if the same lab product or an alternative is used in the 5 most similar protocols
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Purified CD4+/CD8+ or OT-I/-II T cells were labelled with carboxyfluorescein diacetate succinimidyl ester (CFSE) (Becton Dickinson [BD], Franklin Lakes, New Jersey) and stimulated with 0.5µg/mL plate-coated anti-CD3 (clone 145-2C11; Biolegend, San Diego, California) and 1µg/mL soluble anti-CD28 (clone 37.51; Biolegend), or 10µg/mL OVA-derived peptides (AnaSpec, Fremont, California) for 3 days, respectively. Non-mitogenic IL-7 (5ng/mL) served as a negative control (PeproTech, Cranbury, New Jersey). T cells were treated with 0.125-1μM CM-272. Cells were collected for flow cytometry at 72 hours and IFN-γ was measured in culture supernatants at 48 hours. MO4 cells were co-cultured for 48 hours with OT-I TCR-engineered T cells at 1:2 effector/target ratio, while exposed to 0.125-0.5μM CM-272, before IFN-γ measurement.
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