Figure 7. GEM T cells in polyclonal populations ex vivo.

(a) Equal numbers of PBMC were sorted based on staining with antibodies against CD4 and TRAV1-2 and tested using K562 cells transfected with CD1b and CD1c in the absence or presence of GMM or MA. The experiment was performed twice in donors BB12 and BB2, and three times in donor C58, with similar results. (b) PBMC of subject C58 were stained with α-CD3, GMM-loaded CD1b tetramer and α-TRAV1-2. This experiment was performed twice with similar results. (c) The cDNA of 4000 sorted Tet⁺ and Tet⁻ cells was used in a PCR with a TRAV1-2 and TRAJ9-specific primer pair. Cell sorting was performed once, and PCR was performed twice for each donor. (d) Quantity of TRAV1-2–TRAJ9 junctions relative to constant α chain was determined in cDNA from sorted CD4⁺TRAV1-2⁺ PBMC from blood bank donors and tuberculosis patients. Deep sequencing of the TCR repertoire of BB36 and BB38 revealed that 0.48 % and 0.90 % of the TRAV1-2⁺CD4+ T cells, or 0.0024 % and 0.0023 % of the total PBMC population, respectively, expressed typical GEM α chains.