Figure 3. Selection of α–Cbtx V_HH-displayed phage binders.

(A) Polyclonal phage ELISA. The progress of selecting phage-V_HHs specific to α–Cbtx over the three rounds of panning was assessed by doing a polyclonal phage ELISA. Eluted phage (amplified; 100 µL of 10¹¹ pfu mL⁻¹) from each round of panning were incubated with immobilized α–Cbtx (1 µg mL⁻¹) and detected with anti-M13 antibody conjugated to horseradish peroxidise. (B) Monoclonal phage ELISA. Forty-six (not all data shown) monoclonal phage from the 3^rd round of panning were randomly selected and their specificities to α–Cbtx were assayed by indirect ELISA with anti-M13 antibody conjugated to horseradish peroxidase. Absorbance values at 450 nm are the mean of triplicates with background (blocked wells with no α–Cbtx) subtracted. SEMs are shown with bars.