Phage Display for Antibody Discovery & Production

Introduction Basics Hybridoma vs Phage Display Application

Introduction to Phage Display Technology

Phage display involves presenting peptides or proteins on the surface of phages. This approach can match up the genetic data to its phenotype and is thus highly effective in detecting particular protein interactions. At the core of phage display is the screening of peptide or antibody libraries for peptides or antibodies that are very strongly specific to target molecules. Phage display was first reported in 1985 by George P Smith, when phages first revealed they could recognize foreign peptides. It had been deployed for antibody display (and to construct antibody libraries) by the early 1990s. These libraries can be generated from various sources, including raw libraries, vaccines, or synthetic oligonucleotides, allowing for a broad range of potential high-affinity binders to be identified. Phage display technology has since become an indispensable tool in multiple fields, such as therapeutic antibody discovery, vaccine design, and synthetic biology.

Phage Display in Antibody Discovery

Process of phage display antibody discovery

Phage display antibody discovery is an iterative process where antibodies with high specificity and affinity for the antigen are sought.

Library construction: Generate a diverse library of phages displaying antibody fragments (scFv, Fab, or VHH). Library sizes range from millions to billions of variants.
Panning: The library is exposed to the immobilized antigen, and phages that bind strongly to the target are retained, while those that do not bind are washed away.
Screening: After multiple rounds of panning, individual phages are isolated and further screened to identify those with the highest binding affinity and specificity.
Characterization: The screened antibodies are further characterized using techniques such as ELISA, SPR, and immunofluorescence.

Process of phage display antibody discovery Fig. 1 Phage display methodology.^{1, 3}

Classification of phage display libraries

A phage display library is a collection of phages, each of which displays a different peptide or antibody fragment on its surface.

Natural library: an antibody library constructed using lymphocytes that have not been immunized.
Immune library: an antibody library constructed using B cells, tissues, etc. of animals or humans that have been immunized with antigens.
Semisynthetic library: an antibody library constructed by combining a portion of artificially synthesized sequences with another portion of natural sequences.
Synthetic library: an antibody library in which the variable region sequences of antibodies are all artificially synthesized.

Types of antibody phage display formats

Single-chain variable fragment (scFv): A fusion of the variable regions of the heavy and light chains of an antibody, connected by a short peptide linker. ScFvs are small, flexible, and easy to express on the surface of phage.
Fragment antigen binding (Fab): A larger fragment that includes the variable regions of the heavy and light chains and a small portion of the constant region.
Single domain antibody (sdAb): Also known as VHH, they are derived from the variable region of heavy chain antibodies found in camelids. SdAbs are smaller and more stable than traditional antibodies.

Hybridoma vs Phage Display

Both hybridoma and phage display technologies are pivotal in the production of monoclonal antibodies. Hybridoma technology, first developed in the 1970s, involves the fusion of B cells with myeloma cells to generate hybridomas. These hybridomas can be cultured indefinitely and are capable of producing antibodies.

	Phage Display Technology	Hybridoma Technology
Mechanism	In vitro technique using bacteriophages to display peptides or proteins	In vivo technique using hybrid cell lines (fusion of B cells and myeloma cells)
Library Size	Create libraries with billions of variants	Typically smaller, with fewer variants
Target Diversity	Naïve, immunized, semi-synthetic or synthetic libraries	Antibodies produced by immune system of animals
Applications	Antibody discovery, diagnostics, therapeutic applications	Monoclonal antibody production for therapeutics
Advantages	Large-scale production Fast turnaround Easy to control the screening process Easy to screen a large number of different clones Can directly screen human libraries Can be used to screen toxic antigens No immunogenicity issues for naïve libraries No clone viability issues Direct access to sequences No animal use for naïve libraries	Large-scale production High yield High specificity High sensitivity Low cost
Disadvantages	More expensive Binders may have lower affinity More difficult to use	Long generation time May Need humanization

Creative Biolabs will offer customized antibody phage display services to prepare your monoclonal antibodies as fast as possible. We also have a wide range of premade antibody libraries, so we can provide multiple formats to suit a variety of applications.

Applications of Phage Display Antibody Discovery

High-throughput screening: Phage display facilitates rapid screening of large antibody libraries against multiple targets simultaneously.
Development of multispecific antibodies: The technology allows for the creation of multispecific antibodies that can bind to multiple targets.
Antibody engineering: Phage display can be used to engineer antibodies with desired properties, such as increased stability, reduced immunogenicity, and improved pharmacokinetics.
Targeting difficult antigens: Phage display is particularly effective in generating antibodies against challenging targets, including membrane proteins and non-protein antigens such as small molecules and carbohydrates. This versatility stems from its ability to screen large libraries.
Therapeutic applications: Phage display technology has played an important role in the discovery and development of monoclonal antibodies (mAbs) for the treatment of various diseases, including cancer and autoimmune diseases. To date, several approved mAbs have been derived through phage display technology.

Applications of Phage Display Antibody Discovery Fig. 2 The overview of phage display technology.^{2, 3}

References

Almagro, Juan C., et al. "Phage display libraries for antibody therapeutic discovery and development." Antibodies 8.3 (2019): 44.
Zhao, Hui, et al. "Phage display-derived peptides and antibodies for bacterial infectious diseases therapy and diagnosis." Molecules 28.6 (2023): 2621.
Under Open Access license CC BY 4.0, without modification.

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