In vitro assessment of the antitumor activity of CEA-specific T cells. (A) Purified CD8⁺ cells from transplant-recipient patients were sensitized in vitro and then tested in ELISPOT for IFN-γ secretion, after a minimum of 3 stimulation cycles (numbers in parentheses indicate the number of cycles after which the ELISPOT assay was performed). T cells (2 × 10⁴) were plated in the ELISPOT plate either alone (CD8⁺ only) or together with 5 × 10³ CRC cells, as indicated. (B) Unselected PBMCs were similarly tested, but without any prior in vitro stimulation (patient no. 11). PBMCs (1 × 10⁵) were plated in the ELISPOT plate either alone or together with 2.5 × 10⁴ peptide-pulsed T2 cells (CMV-pp65, CEA-Cap1-6D, or Mart1-A27L) or 5 × 10³ colon cancer cells, as indicated. Results represent mean ± SD of 5 replicates. ^*P < .05 as determined by a 2-tailed t test for unpaired samples. SW403 and HT29 are 2 CRC cell lines: although both cell lines express CEA, only SW403 expresses HLA-A^*0201.