Multicolor staining of R3-specific T lymphocytes. CD8⁺ lymphocytes from an AML patient were subjected to one round of stimulation with autologous CD8 APCs in the presence (B) or absence (A) of the R3 peptide or an irrelevant MAGE3-derived peptide (C) as described in “Materials and methods.” A difference in frequency of R3-specific T cells could be noted. To further characterize R3-specific CD8⁺ T cells, lymphocytes were gated (D; gate R1) and stained with HLA-A2/R3 tetramers and anti-CD8*PerCP (E; gate R2). Lymphocytes from gate R2 were analyzed for their counterstaining of CCR7 and CD45RA (F). Most (68%) were revealed to be CD8⁺ HLA-A2/R3 tetramer⁺ CCR7-CD45RA⁺ effector T cells. The figure displays representative results. SSC indicates side scatter; FSC, forward scatter; APC, allophycocyanin (panel F only).