We use an error-prone PCR integrated DNA-shuffling approach to mutate mainly CDR regions during sub-library construction. If the potential of introducing immunogenic mutations to framework positions is not a concern, we usually use this approach to create mutations at completely random positions across the entire VH and VL fragments. In these cases, the genetic diversity of the sub-library is further increased via passage through our proprietary bacterial mutator strain, CD-affi™.

1. antibody avidity
• Affinity maturation is the process to improve antibody affinity for an antigen. In vivo, natural affinity maturation by
the immune system takes place by somatic hypermutation and clonal selection. In vitro, in the laboratory affinity
maturation, can be obtained by mutation and selection.
• Creative Biolabs has gained extensive experience in antibody affinity maturation. We usually take scFv as the
antibody format in affinity maturation. Also, a monovalent display phagemid system is used to reduce the avidity
effects during antigen-binding screening. We also provide affinity maturation services for single domain antibodies.
Two methods, untargeted mutagenesis and oligonucleotide-directed mutagenesis, are employed to construct
random or defined sub-libraries to introduce a large number of mutants of the original antibody. Antibody binders
of higher affinity are then selected by increasing the screening stringency. By constructing a series of sub-libraries
of a scFv/Fab antibody, our proprietary protocol allows increase of the affinity of the scFv antibodies from 10 -9 to
10 -10. We have successfully obtained a scFv antibody that has an extremely high affinity of 10 -12, whose binding
to the antigen is essentially irreversible.

2. • Untargeted Mutagenesis
• Antibody Affinity Maturation
• We use an error-prone PCR integrated DNA-shuffling approach to
mutate mainly CDR regions during sub-library construction. If the
potential of introducing immunogenic mutations to framework
positions is not a concern, we usually use this approach to create
mutations at completely random positions across the entire VH and
VL fragments. In these cases, the genetic diversity of the sub-library is
further increased via passage through our proprietary bacterial
mutator strain, CD-affi™.

3. • Oligonucleotide-directed Mutagenesis
• If the structure of the antibody/antigen complex is available or modeling the
structure of the antibody/antigen is possible, certain positions can be
randomized at a defined diversity (such as full randomization with all 20
amino acids or biased randomization with selected amino acids at fixed
percentages) to improve the affinity. We are able to create any sub-libraries to
incorporate the defined mutations using trimer codon technology. Most of the
time, we need study the AA sequences of the antibody to find out the
conserved sequences (in comparison with the germ-line and antibody
subfamily sequences). We may then introduce mutations to the positions in
the frame work regions that are not conserved. Supposedly, these regions will
be antigen-specific and change in these regions may not increase
immunogenicity.