text: Isolation and culture of primary CD8+ T cells.
  CD8+ T cells from 4-8-wk-old Tcra-/- x P14 TCR transgenic (Taconic), C57BL/6J WT, or Tbx21-/- (The Jackson Laboratory) mice were purified (>95% purity) by negative selection (Invitrogen) from pooled spleen and lymph node cells. CD8+ T cells from Runx3-/- mice on the ICR background were purified by positive selection (Miltenyi Biotec). All mice were maintained in specific pathogen-free barrier facilities and used according to protocols approved by the Immune Disease Institute and the Harvard Medical School Animal Care and Use Committees. For stimulation, purified CD8+ T cells were cultured at 106 cells/ml (10 ml) in T25 flasks coated with 1 mug/ml each of anti-CD3 (clone 2C11) and anti-CD28 (clone 37.51) by pretreatment with 300 mug/ml goat anti-hamster IgG. After 48 h, cells were removed from the TCR stimulation and recultured at a concentration of 5 x 105 cells/ml in media supplemented with 100 U/ml rhIL-2. Every 24 h, viable cells were counted and readjusted to 5 x 105 cells/ml with fresh media containing the corresponding amount of rhIL-2. 
  