Chen, J.; Zhu, X.; Huo, J.; Wu, S.; Zhou, T.; Cheng, C.; Dong, H.; Li, Y.; Chen, Y.; Dong, X.

CD4+ T cells play a pivotal role in adaptive immunity, recognizing peptide antigens presented by MHC II molecules during infections and tumor development. Identifying immunodominant MHC II epitopes is essential for understanding CD4+ T cell responses; however, current methods such as mass spectrometry suffer from low sensitivity and throughput, while computational algorithms show variable accuracy. To overcome these challenges, we developed EliteMHCII, a high-throughput immunopeptidome profiling platform that identifies antigen-derived MHC II epitopes and measures peptide binding affinity across 24 globally common MHC II alleles. Using EliteMHCII, we assessed the immunodominant epitopes of the SARS-CoV-2 RBD protein. Validation in vaccinated individuals and humanized mouse models revealed a strong correlation between high-affinity peptides and robust CD4+ T cell responses, while low-affinity peptides failed to elicit responses. Therefore, our immunopeptidome profiling platform, EliteMHCII, serves as a rapid, high throughput, feasible platform for CD4+ T cell epitope discovery at a global populational level in the context of infectious diseases and cancer immunotherapy.