Fig. 2: 10E8-UCA^H B cells are primed and recruited to GCs.

a, Affinities of human 10E8-UCA^H/UCA^L or 10E8-UCA^H/mouse^L Fabs to 10E8-GT9.2 12mer in a nanoparticle SPR assay. Symbols represent individual antibodies, and the line denotes the median 10E8-UCA^H/mouse^L value. b, Representative flow cytometry plots of CD95^hiCD38^loB220⁺ GC B cells and their distribution into CD45.2⁺ and CD45.1⁺ B cells at day 7 p.i. in CD45.1 wild-type mice transferred with 50,000 CD45.2⁺ 10E8-UCA^H B cells (to reach a frequency of 4:10⁵ CD45.2⁺ 10E8-UCA^H B cells) 1 day before immunization with 50 µg of 10E8-GT9 12mer or 10E8-GT9-KO and Ribi adjuvant. c, Representative flow cytometry plots showing the percentage of CD45.2⁺10E8-GT9⁺⁺ (top) CD45.1⁺10E8-GT9⁺⁺ (bottom) GC B cells at day 7 p.i. with 50 µg of 10E8-GT9.2 12mer in recipients as in b. d, Ribbon diagrams showing the modifications in immunogens 10E8-GT9.4 and 10E8-GT9.6 derived through the reduction of off-target effects (10E8-GT9.4) plus the addition of the PADRE T cell help epitope (10E8-GT9.6). Mutation T95A (red) was removed, as candidates containing this mutation failed to express as 12mers. e,f, Representative flow cytometry plots of CD45.2⁺ GC B cells and CD45.1⁺ GC B cells (e) and quantification of GC B cells and CD45.2⁺ GC B cells (f) at day 7 p.i. in CD45.1 wild-type mice transferred with CD45.2⁺ 10E8-UCA^H B cells (frequency of 6:10⁵ CD45.2⁺ 10E8-UCA^H B cells) and immunized 1 day later with 50 µg of 10E8-GT9.2 12mer, 10E8-GT9.3 12mer, 10E8-GT9.4 12mer, 10E8-GT9.5 12mer, 10E8-GT9.6 12mer or 10E8-GT9-KO 12mer with Ribi adjuvant. Error bars indicate mean ± s.d. from mice in each group (n = 6). Data from one representative experiment are presented, and two independent experiments were performed.