Figure 3: CD8⁺ T cells specific to highly conserved SARS-CoV-2 epitopes detected in COVID-19 patients and unexposed healthy individuals:

(A) Experimental design: PBMCs from HLA-A*02:01 positive COVID-19 patients (n = 30) (B) and controls unexposed healthy individuals (n = 10) (C) were isolated and stimulated overnight with 10 μM of each of the 27 SARS-CoV-2-derived CD8⁺ T cell epitopes. The number of IFN-γ-producing cells were quantified using ELISpot assay (B, C and D). Dotted lines represent threshold to evaluate the relative magnitude of the response: a mean SFCs between 25 and 50 correspond to a medium/intermediate response whereas a strong response is defined for a mean SFCs > 50. PBMCs from HLA-A*02:01 positive COVID-19 patients (E) were further stimulated for an additional 5 hours in the presence of mAbs specific to CD107^a and CD107^b, and Golgi-plug and Golgi-stop. Tetramers specific to Spike epitopes, CD107^a/b and CD69 and TNF-α expression were then measured by FACS. Representative FACS plot showing the frequencies of Tetramer⁺CD8⁺ T cells, CD107^a/b+CD8⁺ T cells, CD69⁺CD8⁺ T cells and TNF-α⁺CD8⁺ T cells following priming with a group of 4 Spike CD8⁺ T cell epitope peptides. Average frequencies of tetramer⁺CD8⁺ T cells, CD107^a/b+CD8⁺ T cells, CD69⁺CD8⁺ T cells and TNF-α⁺CD8⁺ T cells.