Naïve CD25−CD4+ T cells isolated from NOD spleens, MLNs, and head and neck LNs (HNLNs) using immunomagnetic negative selection with an EasySep Mouse Naïve CD4+ T cell Isolation Kit (StemCell Technologies, Vancouver, Canada). Naïve T cells were co-cultured with infected BMDCs at a 1:4 or 1:2 BMDC to T cell ratio. BMDCs and T cells in 200 μL were co-cultured in a round-bottomed, 96-well tissue culture plate that had been coated with anti-CD3 mAb (clone 17A2; Invitrogen), and cells were co-stimulated with soluble anti-CD28 mAb (clone 37.51, Invitrogen). Co-cultures of 1:4 BMDCs to T cells were incubated at 37 °C for 48 h or 1:2 BMDCs to T cells for 5 days. After which culture supernatants were collected and stored at −80 °C until cytokine ELISA analyses could be performed.
Soluble anti cd28 mab clone 37
Manufactured by Thermo Fisher Scientific
Sourced in United States
Soluble anti-CD28 mAb (clone 37.51) is a monoclonal antibody that binds to the CD28 receptor expressed on the surface of T cells. This product is intended for research use only.
Automatically generated - may contain errors
Lab products found in correlation
2 protocols using soluble anti cd28 mab clone 37
1
Naïve T Cell Activation by Infected BMDCs
2
Lymphocyte Stimulation and Cytokine Analysis
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Single cell suspensions from aseptically removed the head and neck lymph nodes (HNLNs), mesenteric LNs (MLNs), and spleens were prepared as previously described [35 (link)]. Lymphocytes were cultured in a complete medium: RPMI 1640 with 2 mM l -glutamine (Genesee Scientific, El Cajon, CA) containing 10% fetal bovine serum (Atlanta Biologicals, Oakwood, Georgia), plus supplements (Invitrogen, Carlsbad, CA), 100 U/mL penicillin, 100 μg/mL streptomycin, 1 mM sodium pyruvate, and 0.1 mM nonessential amino acids. Lymphocytes were cultured at 106 cells/well in 96-well, round-bottomed tissue culture plates (Millipore, Billerica, MA) coated with 5 μg/mL anti-CD3 mAb (clone 17A2; Invitrogen, Carlsbad, CA, USA) plus 2.5 μg/mL of soluble anti-CD28 mAb (clone 37.51; Invitrogen) was stimulated for 48 h at 37 °C. Lymphocytes were stimulated in triplicate for 2 days for flow cytometry analysis or for 4 days for collection of cell culture supernatants, which were then stored at − 20 °C until assayed by cytokine-specific ELISAs.
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