All primary murine CD4+ T cells came from homozygous OT-II TCR transgenic mice (Taconic). The following antibodies were used in this study: biotinylated anti–human-CD25 (clone BC96), biotinylated anti-CD3ε (clone 145-2C11), biotinylated anti-CD43 (clone 1B11), and anti-CD28 (clone 37.51) from BioLegend, and anti-CD3ε (clone 145-2C11) from Bio X Cell. Biotinylated I-Ab presenting OVA(323–339) was obtained from the National Institutes of Health Tetramer Facility for pMHC studies. Ovalbumin peptide 323–339 was obtained from AnaSpec, and recombinant human interleukin-2 was obtained from Prometheus. Latrunculin A was obtained from Cayman Chemical, ML-7 from Santa Cruz Biotechnology, Inc., and 2-APB from Abcam.
Anti cd3ε clone 145 2c11
Manufactured by BioXCell
Sourced in United States
Anti-CD3ε (clone 145-2C11) is a monoclonal antibody that binds to the CD3 epsilon chain of the T cell receptor complex. It is commonly used in research applications for the activation and expansion of T cells.
Automatically generated - may contain errors
Lab products found in correlation
Celltrace violet, by Thermo Fisher Scientific (2 mentions)
2 apb, by Abcam (1 mentions)
Anti cd28 clone 37, by BioLegend (1 mentions)
Latrunculin a, by Cayman Chemical (1 mentions)
Ovalbumin peptide 323 339, by AnaSpec (1 mentions)
Anti cd28 clone 37, by BioXCell (1 mentions)
Anti cd3ε, by BioXCell (1 mentions)
Fetal bovine serum (fbs), by Atlanta Biologicals (1 mentions)
Interleukin 2 (il 2), by R&D Systems (1 mentions)
Celltrace cfse, by Thermo Fisher Scientific (1 mentions)
3 protocols using anti cd3ε clone 145 2c11
1
Murine CD4+ T Cell Activation
2
T Cell Activation and TGF-beta Modulation
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All cells were cultured in RPMI supplemented with 10% heat-inactivated Fetal Bovine Serum (Atlanta Biologicals, Lawrenceville, GA, USA), 2mM L-glutamine (Mediatech, Manassas, VA, USA), and 1% penicillin/streptomycin (Mediatech, Manassas, VA, USA). CD8 T cells were activated with 100 ng/mL OVA257 peptide (SIINFEKL, New England Peptide, Gardner, MA, USA), 1 µg/mL anti-CD3ε (clone 145-2C11, Bio X Cell, West Lebanon, NH, USA), or 1 µg/mL anti-CD3ε and 5 µg/mL anti-CD28 (clone 37.51, Bio X Cell) for 5 days in media supplemented with 30 U/mL IL-2 (R&D Systems, Minneapolis, MN, USA). When noted, cells were incubated with TGFβ1 (EMD Chemicals, Inc., Gibbstown, NJ, USA) at a final concentration of 20 ng/mL.
3
Murine T Cell Purification and Proliferation
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CD4+ T cells were purified from murine spleen and LNs by EasySep immunomagnetic negative selection (StemCell Technologies, 19852). Unless otherwise specified, T cells were stimulated with 8 µg/ml plate-bound anti-CD3ε (clone 145-2C11 Bio X Cell, BE0001-1) in media supplemented with 2 µg/ml anti-CD28 (clone 37.51 Bio X Cell, BE0015-1). For proliferation, T cells were incubated with 5 µM CellTrace CFSE (Thermo Fisher Scientific, C34554) or 5 µM CellTrace Violet (Thermo Fisher Scientific, C34517) in PBS + 0.1% BSA for 13 min at 37°C. T cells were then washed thrice with T cell media before (re)stimulation.
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