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Phage Display .net
Thanks for visiting our website. Phage Display .net reviews the phage display technology and its applications, offering phage display related services.
Vrisko Limited is a company specialized in providing custom services in biotechnology. We provide a large number of phage display related services including screenning, affinity maturation, antibody humanization, scFv-, Fab-, and full-size IgG production. In addition, we can also provide custom mouse and rat monoclonal antibodies, protein expression in different expression systems (e.g. bacterial, mammalian cell, yeast, etc), protein arrays, and much more. We are collaborating with companies that are leading services providers in the biotechnological field to bring all these services to our customers.
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. The technology consists in expressing peptides, proteins or antibody fragments at the surface of phage particles (Smith, 1985; Winter et al., 1994; Kay and Hoess,1996). The purpose is to screen for proteins with specific properties selected from libraries of DNA variants expressed into populations of protein variants on phages surfaces.
What is a phage or bacteriophage?
Phages are viruses that infect bacterial cells. Many of the vectors used in recombinant DNA research are phages that infect the standard recombinant DNA host: the bacterium Escherichia coli. There are several types of phage that have been used as vehicles for phage display including Ff filamentous phage, Lambda and T7. Each of these has advantages and disadvantages with respect to each particular application. The Ff phage family (M13 and its close relatives fd and fl) are excellent cloning vehicles because their size is not constrained by the DNA contained within them. The insertion of foreign sequences within their genome is accommodated simply by the assembly of longer phage particles.
History of phage display
Phage display technology was first introduced in 1985 by George Smith. It was used as an expression vector for presenting a foreign amino acid sequence accessible to binding an antibody. Since then, a large number of phage displayed peptide and protein libraries have been constructed, leading to various techniques for screening such libraries. Phage display technology has had a positive effect on works done in the fields of immunology, cell biology, pharmacology, and drug discovery.
Technical description of phage display
The starting point is usually a library of peptides, proteins or antibodies, which can be 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, to varying degrees. The choice of pVIII or pIII is related to the choice of the type of display, either polyvalent or monovalent display.
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 (see screening of phage display libraries). It can be helpful for preservation of unstable hybridoma clones. The technique is useful 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. It can facilitate the identification of peptides with applications in the prevention of diseases, for example, peptides of interest from a phage display peptide library can be captured with monoclonal antibodies against antigens from pathogens. Also phage particles displaying antigens on their surface can be utilized as vaccine like the antigens from Plasmodium falciparum (J Biol Chem 1988;263:4318-22).One of the most important application of phage display is to generate monoclonal antibodies and improve antibody affinity by affinity maduration.
Advantages of phage display
Phage display is a system for large scale study and selection of proteins based on their binding affinity and specificity. One advantage of phage display is the enormous diversity of variant proteins that can be represented. For example, phage display antibody libraries with diversities as high as 10e10 are routinely constructed. Phage display is highly flexible and selection may be performed in vivo or in vitro. In vitro selection enables phage displayed proteins to be screened not only against a wide range of biological targets but also inorganic ones. Phage display screening formats can be readily modified to manipulate selection conditions and stringencies. Phage display provides a means of rapidly screening large numbers of proteins against potential binding partners. As a rough guide, billions of clones can be screened within a week using phage display.
Phage display antibodies
One of the major advantages of phage display antibodies compared with standard hybridoma technology is that the selection of specific scFv/Fab fragments to a particular antigen 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. Many different methods have been described for antibody selection (e.g. yeast, bacterial or ribosomal display, and picking from arrays), however, antibody phage display has developed into the strongest of these methods. Antibody phage display has great potentials, in addition to extend our capacity to generate antibodies, it provides us with the possibility of direct functional analysis of epitopes.
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 libraries of phages expressing on their surfaces proteins from microorganisms. The purpose of these libraries is to select proteins with a specific function or affinity to another molecule of interest. There is a system called Shotgun Phage display which consists in displaying proteins or domains from microorganisms in a multivalent form using pVIII -fusion. In this system, the genomic DNA is digested and cloned randomly in fusion with the gene of pVIII of the phage capsid (Comb Chem High Throughput Screen. 2001 Apr; 4(2):135-43.). There are other systems of phage display which used the protein pIII to display proteins or domains. Phage display antibodies can be selected from antibodies libraries which have been the main use of the system Phage display. In the antibodies libraries, the phage display antibodies are not expressed as complete antibodies, but as Fab or scFv fragments.
Phage display library of antibodies
The genomic information coding for antibody variable domains 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 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 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 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.
Peptide library
We provide service of screening of peptide library based on phage display system. Several types of peptide libraries are available including 10-mer phage display peptide library, 16-mer phage display peptide library and 20-mer phage display peptide library. We can perform the screening using in-house made peptide library or commercially available peptide library. In addition, we have pIII and pVIII fused peptide libraries.
Find a peptide for your target using our phage display peptide library service – Online inquiry
Peptide library
We provide service of screening of peptide library based on phage display system. Several types of peptide libraries are available including 10-mer phage display peptide library, 16-mer phage display peptide library and 20-mer phage display peptide library. We can perform the screening using in-house made peptide library or commercially available peptide library. In addition, we have pIII and pVIII fused peptide libraries.
Find a peptide for your target using our phage display peptide library service – Online inquiry
Production of monoclonal antibodies by hybridoma technology
In addition of phage display service, we provide monoclonal antibodies services using the hybridoma technology. Visit our website of antibody production to review our list of services of monoclonal antibodies at www.monoclonal-antibodies.net.
Scientific papers
Phage display. George P. Smith and Valery A. Petrenko. Chem. Rev. 1997, 97, 391-410.
Phage Display: A Laboratory Manual By Carlos F Barbass, III, Dennis R. Burton, Jamie K. Scott, Gregg J. Silverman.Concepts in antibody phage display by S Carmen – 2002.Phage
Display Derived Therapeutic Antibodies. Holger Thie et. al. Current Pharmaceutical Biotechnology, 2008, 9, 439-446.
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