Measuring T Cell IFN-γ Response to SARS-CoV-2 Peptides

The peptide pools spanning the entire S/N/E/M protein as consecutive 15-mers overlapping by 10 amino acids were synthesized by Scilight Biotechnology, LLC. Approximately 2.5 mg of each purified peptide in the peptide pool was present per vial. The experiment was conducted as described previously [18 (link)]. Briefly, 96-well plates (BD ELISPOT Set, USA) were coated with anti–IFN-γ capture Ab and incubated overnight at 4 °C. The plates were blocked with the complete culture medium after washing three times. Splenocytes were harvested after the mice were euthanized on days 35 and 120 Fresh single-cell suspensions from each group were plated at 5 × 106 per well, and peptides were added. The plates were then incubated at 37 °C in 5% CO₂ for 22 h and detected using an ELISpot plate reader (Biosys, So. Pasadena, CA, USA). A spot-forming unit (SFU) represents a T cell-secreting IFN-γ.