Phage Display technology review
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Our services include screenning of phage libraries (peptide libraries, protein libraries and antibody libraries), affinity maturation of phage display antibodies, antibody humanization, antibody production, and much more. For more information
What is phage display? Phage display is a powerful tool for selecting peptides, proteins, or antibodies with specific binding properties from a large number of variants in phage libraries.
Description of phage Display technology
The starting point is usually a phage library. A phage library can be a peptide library, protein library or antibody library, which are obtained commercially or constructed in-house. The construction of a phage display library is accomplished by inserting DNA fragments into phage or phagemid genomes which proteins are expressed on the phage coat. This creates a direct physical link between the DNA sequences and their encoding proteins. All five capsid proteins from the phage (pIII, pVI, pVII, pVIII and pIX) have been used to display proteins, peptides or antibodies in phage libraries. The choice of pVIII or pIII in phage libraries is related to the choice of the type of display, either polyvalent or monovalent display.
Phage Display antibodies Recombinant antibodies (e.g scFv and Fab) can be selected from a phage display antibody library. One of the major advantages of phage display antibodies compared with monoclonal antibodies obtained by the standard hybridoma technology is that their selection can be done within a couple of weeks. The starting point is usually an antibody library, of either naive or immune origin, comprising a population of, ideally, 10e9–10e11 clones. After usually three to five rounds of selection, the population is enriched for a high percentage of antibody fragments specific for the target antigen.
Phage display libraries
There are several types of phage display libraries including peptide libraries, protein libraries and antibody libraries. For example, peptides can be selected from peptide libraries, which commonly expressed variants of peptides of 7 to 20 amino acids in fusion with the protein pIII or pVIII of the phages. Pre-made or commercial peptide libraries are available for screening service in our website. Proteins of interest can be selected from a protein library like pre-made phage libraries expressing on their surfaces proteins from microorganisms. The purpose of protein libraries is to select proteins with a specific function or affinity to another molecule of interest such as the Shotgun Phage display system (Comb Chem High Throughput Screen. 2001 Apr; 4(2):135-43.). However, the main use of this technology has been the selection of phage display antibodies from antibody libraries. The antibody libraries express the phage display antibodies commonly as scFv or Fab fragments.
To select phage display antibodies it is neccesary to have or construct a phage library that display antibody fragment on the surface of the phage. The genomic information coding for antibody variable domains to construct antibody libraries is usually derived from B cells of immunized or non – immunized donors. The first generation of phage display library was produced by harvesting B cells from immunized animal or patients previously naturally infected or affected by a disease. An antibody repertoire from immunization is generally restricted to generating antibodies against the antigen of the original immunogenic response. The second generation of phage display antibody libraries is thus obtained from naive large repertoires of human antibody fragments which can be recovered from peripheral B-lymphocytes from non-immunized human donors. The V genes from heavy and light chains are amplified, assembled, randomly combined and cloned to encode a combinatorial library of scFv or Fab fragments. With a naive antibody library specific phage display antibodies can be obtained without any previous contact with the antigen. It can be useful to select antibodies against self, non-immunogenic or relatively toxic antigens (Griffiths, et al. 1993; Vaughan et al. 1996). Naïve antibody libraries have the advantage that they can theoretically be used for an unlimited range of antigens. The third generation of antibody libraries is composed by synthetic libraries. Construction of synthetic phage display libraries involves rearranging VH and VL gene segments in vitro and introducing artificial complementarity determining region (CDRs) of varying loop lengths using PCR and random oligonucleotide primers.
Applications of phage display
Phage display can be used for selection of proteins, peptides, or antibodies with affinity and specificity to a molecule or protein of interest from phage libraries. It can be helpful for preservation of unstable hybridoma clones and to identify molecules that can be recognized and internalized by eukaryotic cells. In addition, it can identify epitopes, mimotopes, functional and accessible sites from antigens (Mol Inmunol 1986; 23:709-15), and postransductional modifications in different molecules (Science. 1993; 260:1113-7). Phage display is an excellent tool for designing vaccines.