Figure 5. Antigen-specific expansion of HCAb-engineered B cells.

(a) Diagram of tonsil organoid system. (b) Representative plots of control or J3-edited tonsil B cells, treated with RNPs plus AAV6-J3 at MOI = 5 × 10⁵ vg/cell. (c-e) Tonsil B cells from n = 3–5 donors were edited with RNPs plus AAV6-J3 at MOI = 1–5 × 10⁵ vg/cell, cultured for 2–4 days, and reconstituted with total autologous TMNCs. Cultures were immunized with HIV gp120 plus Adju-Phos or control PE protein plus Alhydrogel and cells were harvested 12 days later for analyses. (c) Representative panel showing increase in gp120⁺ VHH⁺ cells after immunization with gp120 but not PE, measured by flow cytometry, gated on CD19⁺ CD3⁻ B cells as described in Supplementary Fig. 8. (d) Response of tonsil organoid cultures containing control or edited B cells to gp120 or PE immunization. Matching colors indicate samples from the same tonsil donor. (e) Phenotypes of B cells in tonsil organoids containing control or J3-edited B cells were characterized at day 12 by flow cytometry, with or without gp120 immunization, using the gating strategy described in Supplementary Fig. 8. Error bars show mean ± SEM. Statistics in panel (d-e) were calculated by 1-way ANOVA. * p < 0.05, ** p < 0.01.