Figure 3. Recognition of phage-displayed EGFR domain III mutated variants. Phages displaying human EGFR Dom III_+482–514 mutated variants (where each solvent-exposed residue differing between human and mouse EGFR has been replaced by the aa found in the latter) were produced at a 50 ml scale. Phage-displayed wt Dom III_+482–514 was included as a control. Purified phages (10¹² viral particles/ml) were incubated on microtiter plates coated with either anti-EGFR mAbs (nimotuzumab [A] and cetuximab [B]) or the anti-c-myc tag 9E10 mAb. Bound phages were detected with an anti-M13 mAb conjugated to horseradish peroxidase. Normalized reactivity for each variant was estimated by dividing the signal obtained with each mAb by the reference signal (measured with the anti-tag mAb). Relative reactivity (%) was calculated as the ratio between normalized reactivity of each variant and that of wt domain III. Arrows indicate lack of recognition of individual variants by a given anti-EGFR antibody.