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Immunology, IFReC, Osaka Universityhttps://www.ebi.ac.uk/ena/browser/view/PRJNA1033588Homo sapiensSARS-CoV-2 vaccines have been used worldwide to combat COVID-19 pandemic. To elucidate the factors that determine the longevity of spike (S)-specific antibodies, we traced the characteristics of S-specific T cell clonotypes together with their epitopes and anti-S antibody titers before and after BNT162b2 vaccination over time. T cell receptor (TCR) alpha/beta sequences and mRNA expression of the S-responded T cells were investigated using single-cell TCR- and RNA-sequencing. In donors exhibiting sustained anti-S antibody titers (designated as “sustainers”), S-reactive T cell clonotypes detected immediately after 2nd vaccination polarized to follicular helper T (Tfh) cells, which was less obvious in “decliners”. Even before vaccination, S-reactive CD4+ T cell clonotypes did exist, most of which cross-reacted with environmental or symbiotic bacteria. However, these clonotypes contracted after vaccination. Conversely, S-reactive clonotypes dominated after vaccination were undetectable in pre-vaccinated T cell pool, suggesting that highly-responding S-reactive T cells were established by vaccination from rare clonotypes. These results suggest that de novo acquisition of memory Tfh cells upon vaccination contributes to the longevity of anti-S antibody titers. Overall design: Among 8 age- and sex-matched donors who accepted 2 doses of SARS-CoV-2 vaccine BNT162b2, #4, #13, #15 and #17 had declined anti-S antibody titers from 6 weeks (wks) to 24 wks after the 1st vaccination, while #8, #25, #27 and #28 had sustained antibody titers. PBMCs from these 8 donors at 3 wks, 6wks, and 24 wks after the 1st vaccination were collected. PBMCs from donor #4, #8, #13, #15 and #17 before the vaccinations were also collected. PBMCs were stimulated with SARS-CoV-2 S peptide pool, and proliferated T cells were collected and analyzed by scRNA/TCR-seq.ENAfalseT cell responses in SARS-CoV-2 vaccinees with sustained or declined antibody titerT cell responses in SARS-CoV-2 vaccinees with sustained or declined antibody titer2025-09-242023-12-20PRJNA1033588GSE2465359606