SARS-CoV-2 spike protein and CD4-RE-specific CD4+ T cell reactivity. Mild (n = 6, blue), moderate (n = 6, yellow), and severe (n = 14, red) acute COVID-19 donors were assessed for T cell responses to SARS-CoV-2 S and non-Spike proteins. PBMCs from these donors were assayed with the S peptide pool of overlapping 15-mers by 10 and the CD4-RE pool of experimentally defined non-spike epitopes (Yu et al., Cell Host & Microbe 2022). (A) The magnitude for AIM+ CD4+ T cells specific for spike (circles) and CD4-RE (triangles) is shown for mild/moderate and severe COVID-19 donors. The dotted line indicates the LOS for the AIM assay, while the y-axis starts at the LOD. The frequency of positivity is indicated in pie charts for S and CD4-RE for each disease severity. (B) The ratios of CD4-RE to S AIM+ CD4+ responses are shown for responding donors (mild n = 4, moderate n = 2, and severe n = 3). Mann–Whitney or Wilcoxon tests were applied and p-values are shown for significant values. * p < 0.05.