Extended Data Fig. 4: Identification of mimotopes specific for anti-mouse CD19 and anti-ALK antibodies.
(a-d) Identification of a mimotope for anti-mouse CD19 mAb 1D3. (a) Flow cytometry scatter plots illustrating successful identification of yeast cells binding to 1D3IgG at 1 μM. Two yeast populations, P1 and P2, were identified. (b) Top-ranked 1D3 mimotopes. Shown in the table are individual mimotopes and mimotope fusions. (c) ELISA showing the binding of 1D3IgG to chemically synthesized mimotopes in B. A one site-specific binding model was used for assessing the apparent binding affinity. (d) Mouse IFN-γ secretion from anti-mouse CD19 CAR-T cells co-cultured with target cells labeled with 100 nM of amph-mimotopes or free mimotope peptides. Error bars show mean ± s.d. with three replicates. **, p < 0.01 by one-way ANOVA with Tukey’s post-test. (e-f) Identification of a mimotope for ALK-specific mAb ALK123. (e) Flow cytometry plots showing the successful identification of yeast cells binding to ALK123IgG at 500 nM. (f) Binding of ALK123IgG to select yeast clones. Amino acid sequences were shown for each yeast clone. A one site-specific binding model was used for assessing the apparent binding affinity. (g) ELISA showing the binding of ALK123IgG to chemically synthesized ALK123 mimotope E4 (ALK123E4) (n = 3). Error bars show mean ± s.d. with three replicates (h) Representative histogram showing ALK123IgG binding to target cells labeled with amph-ALK123E4. (i) Mouse IFN-γ secretion from anti-mouse ALK123 CAR-T cells co-cultured with target cells labeled with 100 nM or 1 μM of amph-ALK123E4 or NH-SY5Y (ALK+) cells (****, p < 0.0001). Error bars show mean ± s.d. with three replicates. ***, p < 0.001; ****, p < 0.0001; ns, non-significant by one-way ANOVA with Turkey’s post-test.
