Engineering Antibody Affinity

Display technologies have been widely used to increase antibody affinity to values not typically achievable using hybridoma technology. Mutations are introduced into the DNA of an antigen specific antibody fragment either randomly using, for example, error prone PCRor specifically using spiked oligonucleotides. The mutated antibody fragment DNA is then used to create a display library either in phage or yeast or by using ribosome display. Higher affinity antibody fragments are selected from lower affinity ones by a variety of different affinity chromatography approaches, including flow cytometry for yeast displayed libraries. For example, we reported increasing the affinity of an HER2 scFv using phage display by more than 1000-fold from 16nM to 13 pM. Others have reported comparable levels of affinity maturation to values down to low picomolar to femtomolar affinities using either phage or yeast display. These in vitro affinity maturation approaches have now become routine in the biotechnology field.