Antibody Engineering: Humanization, Affinity Maturation, and Selection Techniques

De Novo Selection of High-Affinity Antibodies from Synthetic Fab Libraries Displayed on Phage as pIX Fusion Proteins

“…This process is akin to chain shuffling 45,46 and our vector design is amenable to direct introduction of diversity into individual clones or populations without isolation and characterization of primary hits. Almagro and Strohl 51 have compiled examples of antibodies that have undergone in vitro affinity maturation. The compilation includes affinity maturation strategies using varied display platforms diverse methods for library generation such as error-prone PCR, focused saturating mutagenesis at CDR-H3, and CDR walking mutagenesis.…”

The functional repertoire of rabbit antibodies and antibody discovery via next-generation sequencing

“…1 Rabbit antibodies are well known for their high affinity, exquisite specificity and ability to recognize unique epitopes. 2 Rabbits are also evolutionarily more distant from humans than mice and rats, and thus can generate antibodies against epitopes conserved between rodent and human antigens.…”

Application of Antibody Engineering in the Development of Next Generation Antibody-Based Therapeutics

“…The success of humanization, combined with numerous patents protecting the original CDR grafting method (US Patent 5,225,539 to Winter and Jones) and its variations (US Patent 5,693,761 to Queen et al and 5,821,337 to Carter et al) fueled the diversification of humanization methods in the last two decades. Some of the methods developed in the 1990s and 2000s, often called rational methods (Almagro and Strohl 2009), include resurfacing (Padlan 1991), deimmunization (De Groot et al 2006), specificity-determining residues grafting (Tamura et al 2000), superhumanization (Tan et al 2002), human string content optimization (Lazar et al 2007), and germline humanization (Pelat et al 2008). These methods have in common the design of fewer humanized variants to be tested for binding or any other property of interest based on sequence and structural considerations.…”