Mouse naive (CD62Lhi44lo25−) CD4+ T cells were purified by flow cytometry and activated in vitro with 5 µg/ml plate-bound anti-CD3 and 1 µg/ml soluble anti-CD28 antibodies (eBioscience) in RPMI supplemented with 10% FCS, 5 mM β-mercaptoethanol, and antibiotics.