Table 2: Validation runs for the selection and amplification of Melan-A- and MELOE-1- specific T lymphocytes from HLA-A2 melanoma patient-derived PBMC.
Number of stimulated PBMC
Number of sorted cells and fraction of specific T cells
Theoretical number of specific T cells1
Recovered number of sorted specific T cells
Final number of amplified specific T cells and amplification yield3
Average number of cell divisions
Purity of amplified T cells4
Activity of amplified T cells on the cognate peptide5
The theoretical number of specific cells is estimated from the % of tetramer-positive cells in the sorted population. Sorting yields are calculated by dividing the number of rosetted T cells after sorting by the theoretical number of specific T cells in the sorted populations. Amplification factors are estimated from the final number of amplified T cells divided by the number of rosetted T cells seeded for amplification on feeder cells. Purity of sorted and amplified T cells is assessed by tetramer/CD8 double staining. % of TNF producing T cells among tetramer positive cells : reactivity of amplified T cells is assessed by tetramer/TNF double staining, after peptide activation.