PBMCs isolated from 3 donors (CCP4, VR36, and VR58) were expanded for 10 days with S_815-827 or homologous peptides from HCoV-NL63, MERS-CoV, NL63-bat, 229E-bat, and CBC. HIV-1 Nef peptides were included as a specificity control. After culture, CD4⁺ T cells were isolated and TCR Vβ CDR3 sequencing was done to identify antigen-specific memory T cells that expanded in response to relevant antigen (ViraFEST assay). Cross-reactivity was defined by the functional expansion of the same CD4⁺ TCR clonotypes in response to multiple coronavirus peptides. Peptide coculture was done in triplicate. Data are shown as the (%) frequency after culture (y axis) of antigen-specific CD4⁺ T cell clonotypes (z axis) for all peptide pools tested (x axis). Solid colors represent significant clonotypic expansion in response to the indicated antigenic peptide pool(s), whereas translucent colors indicate the clonotype was present at low frequency in the well but did not significantly expand. Gray colors indicate the relevant TCR clonotype was not detected in that well. Different colors indicate different patterns of cross-reactive T cells shown in a key above each figure. Cross-reactive clones for CCP4 (A), VR58 (B), and VR36 (C) are shown, with different patterns of cross-reactive T cells color-coordinated. CCP, COVID-19 convalescent patient; VR, vaccine recipient; NL63-bat, NL63-related bat; 229E-bat, 229E-related bat; CBC, Chaerephon bat coronavirus; HIV, HIV-1 Nef.