(A) Timeline showing the vaccination history of the research subject, with exposures and repertoire sequencing indicated. (B) Phylogenetic trees showing the FluA-151 lineage over 4 years of vaccination and infection, with branch colors corresponding to sequencing time point. At left, the heavy chain phylogeny for FluA-151 is color-coded based on year of vaccination and days postvaccination. At right, the light chain phylogeny for FluA-151 is shown. Paired heavy-light sequences identified by single-cell sequencing are shown with dashes connecting the heavy chain and light chain trees. (C) Binding of TI mAb FluA-151, a clonally related mAb (FluA-151_Sib1), and the inferred unmutated common ancestor of FluA-151 (FluA-151 UCA) to a panel of recombinant HAs. The previously described TI mAb FluA-20 and a recombinant version of the broadly reactive stem mAb MEDI-8852 are shown for comparison. FluA-20 and FluA-151 did not bind measurably to the A/New York/107/2003 HA, which has a 220 loop deletion. (D) ELISA binding of FluA-151 point mutants to HAs from different strains. Points and error bars represent the mean ± SD of technical triplicates. Experiments were repeated twice, with data from one representative experiment shown. (E) FluA-151 protection from weight loss in a sublethal H1N1 challenge model. Mice were passively transferred with either FluA-151 WT (blue), FluA-151 LALA-PG (red), the positive control anti-stem mAb MEDI-8852 (orange), or the negative control anti-dengue mAb 2D22 (purple) 1 day prior to intranasal challenge with a sublethal dose of A/California/04/2009. For weight loss curves, error bars show the SEM. Statistical comparisons between treatment groups were performed using a repeated measurements 2-way ANOVA with Tukey’s correction for multiple comparisons.