Staphylococcus aureus enterotoxin b

The EBV-B cell lines CP50-EBV and LG2-EBV were obtained several years ago in our laboratory by immortalization of blood B cells with EBV, using a published procedure55 (link). Cells were cultured in Iscove's modified Dulbecco's medium (IMDM; Life Technologies–Carlsbad, CA, USA) supplemented with 10% foetal calf serum, 0.24 mM L-asparagine, 0.55 mM L-arginine, 1.5 mM L-glutamine (AAG), 100 U ml⁻¹ penicillin and 100 μg ml⁻¹ streptomycin. Patient-derived tumour ascites and blood cells from haemochromatosis patients were collected after approval by Institutional Review Boards of all collaborating institutions. CD8 TILs and blood CD8 T cells were sorted by a negative selection strategy (Miltenyi Biotec–Bergisch Gladbach, Germany) from the CD2-positive cells selected by rosetting from the ascites using sheep red bloods cells23 (link). In some experiments, freshly isolated blood CD8 T cells were first cultured in IMDM, 10% human serum (HS) and 25 U ml⁻¹ rhIL-2 (Chiron) in the presence of irradiated LG2-EBV cells, which were previously pulsed with 10 ng ml⁻¹ of a cocktail of bacterial sAg: toxic shock syndrome toxin-1, Staphylococcus aureus enterotoxin A and Staphylococcus aureus enterotoxin B (all from Sigma-Aldrich–Saint-Louis, MO, USA). These T-cell lines were used after 1–2 weeks in culture.