Figure 3. Binding of multi-specific zybodies. (A) SPR analysis demonstrated simultaneous binding of the penta-specific zybody, a2H-n1L-TRA-i1H-e1L, to five targets. EGFR-Fc (20 μg/ml) was immobilized on a CM5 sensor chip using the amine coupling procedure described in the Biacore 3000 manual. This was followed by sequential exposure to the zybody (30 μg/ml), ErbB2-Fc (20 μg/ml), IGF-1R (20 μg/ml), Ang2 (2 μg/ml) and integrin αvβ3 (5 μg/ml). Each ligand was injected for 4 min. (B) Summary of direct binding EC₅₀ values for select zybodies and comparitor antibodies determined by ELISA. The full dilution curves are provided in Figure S3.