Extended Data Figure 24:

Characterization of scFvs that bind TcdB using yeast surface display flow cytometry

A) (left) Results of flow cytometry of yeast samples displaying scFv4-C-Myc construct. Each sample was treated with a titration of soluble biotinylated TcdB fragment (1401–1616) (bn-TcdB) concentrations and visualized with anti-C-Myc FITC + SAPE. (right) Percentage of expressing cells which are within the gate increase with bn-TcdB concentration. B) (left) scFvs were designed to bind to the Frizzled-7 epitope and therefore should compete with Frizzled-7. Designed scFvs should not compete with CSPG4, which binds at a different epitope on full-length TcdB. (right) Yeast displaying scFv4-C-Myc were incubated with 1 nM TcdB and either no competitor, 100 nM Frizzled-7, or 100 nM CSPG4. Binding signal decreases when Frizzled-7 is added, supporting that scFv4 binds at the Frizzled-7 epitope. Binding signal does not significantly decrease when CSPG4 is added.