Journal: iScience

Article Title: Integrating complementary approaches reveals antigen-reactive CD4 + T cell states after SARS-CoV-2 vaccination

doi: 10.1016/j.isci.2026.116175

Figure Lengend Snippet: Identification of SARS-CoV-2-specific T cell responses by pHLA class II dextramers (A) UMAP with Leiden clusters (cluster names in UMAP, cluster numbers on the right) of CD4 + T cells enriched for pHLA dextramer-binding ( A) ( n = 4,882 cells). (B) Dot plots show the log-normalized expression of representative genes and centered log-ratio (CLR) transformed expression of surface proteins per cluster. Numbers on the left indicate cluster numbers. (C) Visualization of clone sizes for all cells in the UMAP. (D) Log-normalized expression of selected genes and scores. (E) Visualization of SARS-CoV-2 spike epitope-specific T cells (DPB1∗04:01/S 167 : n = 38) across all HLA-matched donors (CoVa-Adapt donors and HIM; n = 7) and screened time points. (F and G) UMAP visualization (F) and quantified fractional distribution (G) of DPB1∗04:01/S 167 -specific T cells of HLA-matched CoVa-Adapt donors excluding A7 due to breakthrough infection ( n = 5) at individual time points after primary (P), secondary (S), and tertiary (T) vaccination. Colors represent the cluster location of epitope-specific cells at the respective time points. Cells without the indicated epitope-specificity are shown in gray. Time points with no detected epitope-specific cells are indicated as n.d. (H) Similarity network of TCRs identified as reactive in the reverse phenotyping dataset (dark green) and of DPB1∗04:01/S 167 dextramer + TCR clones (orange) together with previously published SARS-CoV-2-specific TCRs (left). Each vertex in the similarity network represents a unique paired αβTCR clonotype, and edges connect vertices with ≤120 TCRdist units. Only clusters containing at least two clonotypes are shown. Colors indicate SARS-CoV-2 epitope-specificity and dataset origin. Clonotypes without assigned specificity in the published dataset are shown in gray. Clone IDs from our own datasets are provided for each vertex, with functionally tested clones highlighted in bold. Clones for which SARS-CoV-2 spike reactivity could be functionally validated are highlighted with a green asterisk, while non-reactive clones are marked by a red asterisk. Two clusters containing our reactive and DPB1∗04:01/S 167 -specific clones, together with spike-annotated published clones, are highlighted (left and right clusters). Sequence motifs of CDR3α and CDR3β regions for the highlighted clusters are shown on the right. Amino acid positions are indicated at the bottom of each plot. For the CDR3α region, the dominantly used V- and J-gene segments are indicated below the motifs. For detailed V- and J-gene segment usage, see I.

Article Snippet: For each cocktail, 1 μL per 5×10 6 cells of HLA-matched dCODE Dextramers (immudex) and 100 μM d-biotin (1/10 of total dextramer volume) were pre-mixed in FACS-buffer to block free binding sites.

Techniques: Binding Assay, Expressing, Transformation Assay, Infection, Clone Assay, Sequencing