Fig. 2.

Nonconservative substitutions in the CDRH3 loop of 4E10 do not significantly alter epitope peptide affinity. (A) Binding of IgG1 4E10 and variants to a biotinylated epitope peptide (bio-29) in a neutravidin capture assay. Antibody–bio-29 association was allowed to reach equilibrium before transferring to a neutravidin-coated 96-well plate for a short incubation period. Neutravidin-captured antibody–bio-29 complexes were quantified using an alkaline-phosphatase conjugated, anti-human, Fc-fragment specific antibody. IgG1 4E10 P, commercially available from Polymun Scientific, was included as a positive control. (B) Half-maximal effective concentration (log EC₅₀) and corresponding SEM for each antibody against bio-29. Values were derived by nonlinear regression analyses from the bio-29 binding curves (GraphPad Prism 5.0 for Windows).