Ifosfamide and BSO have no effect on DC antigen-specific T-cellstimulation. (A) DCs were cultured for 6 days in medium containing GM-CSF and IL-4 alone. TNF-α (800 U/mL) was added from days 6 to 8 of culture. BSO (0.5 mM) was added for 24 hours on day 7, and ifosfamide (100 μM) was added for 90 minutes on day 8. Treated and untreated DCs were then plated at 10⁴/well in 100 μL medium in 96-well round-bottom plates and pulsed with tyrosinase peptide (1-10 μg/mL) for 2 hours. Tyrosinase-specific T cells (2 × 10⁴) were then added in a final volume of 100 μL to each well in medium containing 20% FCS and 100 U/mL IL-2. Control cultures containing non-tyrosinase-pulsed DCs, DCs alone, or CTLs alone were also included. Supernatants (100 μL) were removed from each well after 24 hours and assayed for IFN-γ production using an ELISA kit specific for IFN-γ. Results represent the mean ± SD of triplicate samples from 1 of 3 representative experiments. (B) 4-OH-IF- or BSO-treated DCs were washed, and GSH levels were measured by HPLC. Untreated cells were used as controls. Results are expressed as the percentage of control values and represent the mean ± SD from 3 treatments. ^*P < .05.