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To determine the role of IDO1-associated Trp metabolism in NNK-induced suppression of CD8 + T cells, CD8 + T cells were isolated from the spleen and lymph nodes (LNs) of C57BL/6 mice using PE-CY7-labeled anti-CD8 antibody by flow cytometric cell sorting, labeled by 5-(and-6)-carboxyfluorescein diacetate succinimidyl ester (CFSE), and cultured alone or cocultured with si IDO1 -treated LLC cells (2:1) in RPMI 1640 medium with 10% FBS, 1 ng/ml IL-2 and activation by the precoated 5 g/ml anti-CD3 antibody and 5 g/ml CD28 antibody