What is a phage or bacteriophage?
Phages are viruses that infect bacterial cells. There are several types of phages that have been used as vehicles for phage display including Ff filamentous phage, Lambda and T7. In phage display, the bacteriophages are used to construct different phage libraries as peptide libraries or antibody library
History of phage display
Phage display technology was first introduced in 1985 by George Smith. After that many advantages have been introduced including the way to construct a phage library like antibody library to select phage display antibodies.
Advantages of phage display
Phage display is a system for large scale study and selection of proteins, peptide and antibodies based on their binding affinity and specificity. One advantage of phage display is the enormous diversity of variant proteins that can be represented in a phage library. Phage display provides a means of rapidly screening large numbers of proteins against potential binding partners.

Antibody Affinity Maturation

Review of affinity maturation of antibodies

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

 

 Methods of antibody affinity maturation

There are mainly two methods for antibody affinity maturation: untargeted mutagenesis and oligonucleotide-directed mutagenesis. Untargeted mutagenesis (e.g. chain shuffling) consists in integrating mutations in the CDR regions by error-prone PCR or other methods. Oligonucleotide-directed or site-directed mutagenesis consists in creating mutations at a defined site in a DNA molecule.

 

One of the most popular methods for antibody affinity maturation is chain shuffling. Chain shuffling consists in fixing one of the two chains (VH and VL) and combining with a repertoire of partner chains to yield a secondary phage display library which can be used to search antibodies with superior affinity. This approach mimic the natural “random” mutations taking place in vivo when it is introducing mutations into VH and VL germline genes.

Example of antibody affinity maturation by random mutagenesis. Random point mutations are incorporated in the variable regions using an error prone PCR technique with a DNA polymerase that makes mistakes in the base paring during DNA synthesis that result in the introduction of errors in the newly synthesized complementary DNA strand. The library of variable regions mutants plus fraction of the constant region are displayed on the surfaces of phages, and mutant antibodies with improved affinities are selected by binding to antigen under stringent selection conditions

 

Antibody affinity maturation services

Vrisko Limited offers a variety of Custom Services including Antibody Affinity Maturation services. Ask for information and quotation at info@vrisko.net