Selecting Wnt Receptor-Binding Fabs

The phage-displayed synthetic library F was used to select for Fabs that bound to Wnt receptors, as described (Persson et al., 2013 (link)). Briefly, Fc-tagged ECD protein (R and D Systems) was immobilized on Maxisorp immunoplates (ThermoFisher, catalog number 12-565-135) and used for positive binding selections with library phage pools that were first exposed to similarly immobilized Fc protein to deplete non-specific binders. After 4 rounds of binding selections, clonal phage were prepared and evaluated by phage ELISA (Birtalan et al., 2008 (link)). Clones that displayed at least 10-fold greater signal for binding to antigen compared with Fc were considered to be specific binders that were subjected to further characterization.