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-In vivo use of phage libraries. Initially the phage library is injected in the circulation. Next, phage is allowed to circulate for minutes or hours (in this case when internalized phages are to be recovered). Finally, after deep anesthesia, mice are euthanized and the organs are dissected for phage recovery and peptide sequencing.
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The expression of exogenous peptides on the surface of filamentous bacteriophage was initially described by Smith in 1985. Since his first study, different molecules such as small peptides and antibodies have been displayed on coat proteins of phage, greatly expanding the applications of the technology. The past decade has seen considerable progres...
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The emergence of antibiotic-resistant-bacteria is a serious public health threat, which prompts us to speed up the discovery of novel antibacterial agents. Phage display technology has great potential to screen peptides or antibodies with high binding capacities for a wide range of targets. This property is significant in the rapid search for new a...
Despite the decades of research, cancer causes millions of deaths every year. Therefore, the development of methods for the indication of tumor markers is very relevant. Heat shock proteins (HSPs) are important for oncogenesis and malignant progression and are a marker of cancer development. Here, HSP-specific antibodies were prepared by using a no...
The selection process aims sequential enrichment of phage antibody display library in clones that recognize the target of interest or antigen as the library undergoes successive rounds of selection. In this review, selection methods most commonly used for phage display antibody libraries have been comprehensively described.
Malignant tumor is one of the leading causes of death in human beings. In recent years, bacteriophages (phages), a natural bacterial virus, have been genetically engineered for use as a probe for the detection of antigens that are highly expressed in tumor cells and as an anti-tumor reagent. Furthermore, phages can also be chemically modified and a...
Citations
... The concept of phage display technology was first demonstrated in 1985 by Smith, who accomplished the incorporation of foreign DNA into the chromosome of the M13 phage [17]. This resulted in the fusion of foreign peptides with the G3P coat protein of the M13 phage [3]. ...
... By binding to a phage coat protein, peptides, proteins, and antibody fragments are expressed on the phage surface. Additionally, the introduction of exogenous DNA sequences permits the display of related genes and their products on the phage surface (58,59). Moreover, the protein or peptide maintains its ability to recognize molecular targeted binding sites (60). ...
Cholera, a persistent global public health concern, continues to cause outbreaks in approximately 30 countries and territories this year. The imperative to safeguard water sources and food from Vibrio cholerae, the causative pathogen, remains urgent. The bacterium is mainly disseminated via ingestion of contaminated water or food. Despite the plate method’s gold standard status for detection, its time-consuming nature, taking several days to provide results, remains a challenge. The emergence of novel virulence serotypes raises public health concerns, potentially compromising existing detection methods. Hence, exploiting Vibrio cholerae toxin testing holds promise due to its inherent stability. Immunobiosensors, leveraging antibody specificity and sensitivity, present formidable tools for detecting diverse small molecules, encompassing drugs, hormones, toxins, and environmental pollutants. This review explores cholera toxin detection, highlighting phage display-based nano immunosensors’ potential. Engineered bacteriophages exhibit exceptional cholera toxin affinity, through specific antibody fragments or mimotopes, enabling precise quantification. This innovative approach promises to reshape cholera toxin detection, offering an alternative to animal-derived methods. Harnessing engineered bacteriophages aligns with ethical detection and emphasizes sensitivity and accuracy, a pivotal stride in the evolution of detection strategies. This review primarily introduces recent advancements in phage display-based nano immunosensors for cholera toxin, encompassing technical aspects, current challenges, and future prospects.
... Therefore, a physical link between the phenotype and genotype of the displayed peptide can be observed [15]. Phage display libraries can be constructed containing up to 10 10 different variants simultaneously, which effectively allows affinity screening of the combinatorial peptide library and selection (biopanning) to identify targeted bindingpeptide [16]. Recent studies have proved the phage display system to serve as an efficient tool in the rapid identification of novel peptides against different target receptors through high-throughput screening, such as Tanzeum (glucagon-like peptide-1 receptor agonist) being clinically used to control glycemia in patients with type 2 diabetes mellitus [17], peptide ESCP9 targeted oesophageal squamous carcinoma cells with high specificity and were potent to be developed as an imaging detection probes [18] as well as peptide-based inhibitors against β-lactamase enzyme to overcome antibiotic resistance in bacteria [19]. ...
Cancer is one of the leading causes of mortality worldwide; nearly 10 million people died from it in 2020. The high mortality rate results from the lack of effective screening approaches where early detection cannot be achieved, reducing the chance of early intervention to prevent cancer development. Non-invasive and deep-tissue imaging is useful in cancer diagnosis, contributing to a visual presentation of anatomy and physiology in a rapid and safe manner. Its sensitivity and specificity can be enhanced with the application of targeting ligands with the conjugation of imaging probes. Phage display is a powerful technology to identify antibody- or peptide-based ligands with effective binding specificity against their target receptor. Tumour-targeting peptides exhibit promising results in molecular imaging, but the application is limited to animals only. Modern nanotechnology facilitates the combination of peptides with various nanoparticles due to their superior characteristics, rendering novel strategies in designing more potent imaging probes for cancer diagnosis and targeting therapy. In the end, a myriad of peptide candidates that aimed for different cancers diagnosis and imaging in various forms of research were reviewed.
... Different types of libraries for different groups of biotherapeutics (e.g., scFV or Fabs) could be designed by phage display technology. A detailed discussion of different phage display libraries and production methods is beyond the scope of this review, and can be found in these reviews [41][42][43][44][45][46]. This technology, however, suffers from the method's complexity, the high cost and the fact that it is very time-consuming to generate a library. ...
Antibody fragments are used in the clinic as important therapeutic proteins for treatment of indications where better tissue penetration and less immunogenic molecules are needed. Several expression platforms have been employed for the production of these recombinant proteins, from which E. coli and CHO cell-based systems have emerged as the most promising hosts for higher expression. Because antibody fragments such as Fabs and scFvs are smaller than traditional antibody structures and do not require specific patterns of glycosylation decoration for therapeutic efficacy, it is possible to express them in systems with reduced post-translational modification capacity and high expression yield, for example, in plant and insect cell-based systems. In this review, we describe different bioengineering technologies along with their opportunities and difficulties to manufacture antibody fragments with consideration of stability, efficacy and safety for humans. There is still potential for a new production technology with a view of being simple, fast and cost-effective while maintaining the stability and efficacy of biotherapeutic fragments.
... The direct link between genotype and phenotype makes it easy to express additional peptide sequences on the viral surface. In this way, for example, randomized libraries with a tremendous number of diverse peptide motifs can be generated and screened against almost any target [10][11][12]. Since its introduction to the scientific world in 1985 by George P. Smith, the PSD technique has become an almost universal tool for a constantly growing number of different questions and applications [10,13]. ...
In recent years, the application focus of phage surface display (PSD) technology has been extended to the identification of metal ion-selective peptides. In previous studies, two phage clones—a nickel-binding one with the peptide motif CNAKHHPRCGGG and a cobalt-binding one with the peptide motif CTQMLGQLCGGG—were isolated, and their binding ability to metal-loaded NTA agarose beads was investigated. Here, the free cyclic peptides are characterized by UV/VIS spectroscopy with respect to their binding capacity for the respective target ion and in crossover experiments for the other ion by isothermal titration calorimetry (ITC) in different buffer systems. This revealed differences in selectivity and affinity. The cobalt-specific peptide is very sensitive to different buffers; it has a 20-fold higher affinity for cobalt and nickel under suitable conditions. The nickel-specific peptide binds more moderately and robustly in different buffers but only selectively to nickel.
... 29,30 The phage display method is a fast, simple, accessible and relatively inexpensive technology that can select highaffinity ligands and define relevant mimetic peptides that can be used in diagnostic assays and vaccine systems. [31][32][33][34][35] The technique is an important low-cost approach for the identification of peptides that can be used in different health areas; the results can be quickly defined and readily used for the diagnosis of several human infectious diseases. [35][36][37][38][39] The identification of mimetic peptides of C. trachomatis may be the basis for future methods for rapid diagnosis (point of care), which could be used in medical offices within local infectious disease support clinics, home care testing and screening tests for epidemiological studies, among others. ...
Background:
Chlamydia trachomatis infection is a major public health problem and the most common sexually transmitted infection in the world. Although highly prevalent, 70% to 80% of cases are asymptomatic and undiagnosed.
Purpose:
To overcome some limitations in terms of rapid diagnosis, phage display technology was used to bioprospect peptide mimetics of C. trachomatis immunoreactive and immunogenic antigens to be selected for the production of synthetic peptides.
Methods:
Initially, IgG from 22 individuals with C. trachomatis and 30 negative controls was coupled to G protein magnetic beads. The phage display technique consisted of biopanning, genetic sequencing, bioinformatics analysis and phage ELISA.
Results:
Clones G1, H5, C6 and H7 were selected for testing with individual samples positive and negative for C. trachomatis. Reactions were statistically significant (p < 0.05), with a sensitivity of 90.91, a specificity of 54.55, and AUC values >0.8. One-dimensional analysis with C. trachomatis components indicated that the G1 clone aligned with cell wall-associated hydrolase domain-containing protein, the H5 clone aligned with glycerol-3-phosphate acyltransferase PlsX protein, the C6 clone aligned with a transposase and inactivated derivatives, and the H7 clone aligned with GTP-binding protein. Molecular modeling and three-dimensional analysis indicated the best fit of the four clones with a protein known as chlamydial protease/proteasome-like activity factor (CPAF), an important virulence factor of the bacterium.
Conclusion:
The peptides produced by phage display are related to the metabolic pathways of C. trachomatis, indicating that they can be used to understand the pathogenesis of the infection. Because of their high sensitivity and AUC values, the peptides present considerable potential for use in platforms for screening C. trachomatis infections.
... Since many of the non-antibody-based binders are selected through phage display or a similar display technique, it is appropriate to introduce this approach briefly. Phage display is a molecular biology technique in which large, highly variable sequence libraries are presented as peptides or proteins on filamentous phage surfaces to enable the selection of binders with specificity and high affinity to almost any target, provided the target protein is available in sufficient purity (quality) and amount [71]. To select a binder for a target using phage display, a process referred to as biopanning is performed. ...
There is often a need to isolate proteins from body fluids, such as plasma or serum, prior to further analysis with (targeted) mass spectrometry. Although immunoglobulin or antibody-based binders have been successful in this regard, they possess certain disadvantages, which stimulated the development and validation of alternative, non-antibody-based binders. These binders are based on different protein scaffolds and are often selected and optimized using phage or other display technologies. This review focuses on several non-antibody-based binders in the context of enriching proteins for subsequent liquid chromatography-mass spectrometry (LC-MS) analysis and compares them to antibodies. In addition, we give a brief introduction to approaches for the immobilization of binders. The combination of non-antibody-based binders and targeted mass spectrometry is promising in areas, like regulated bioanalysis of therapeutic proteins or the quantification of biomarkers. However, the rather limited commercial availability of these binders presents a bottleneck that needs to be addressed.
... Exogenous DNA sequences of interest are introduced into a specific location in the phage genome nucleotide sequence, which encodes one of the phage coat proteins (Fig. 1). When phage infection occurs, phage gene expression begins inside the bacterial host, and the inserted peptide/antibody fragment is subsequently displayed on the surface of the phage as a combination product of the relevant genes encoding the coat protein and the cloned sequence (Arap 2005;Aghebati-Maleki et al. 2016). Therefore, if the cloned sequence is randomized, phage display libraries, providing >10 10 variants, can be constructed at the same time and stored for long term as DNA clones, rather than requiring the individual construction of various peptides or antibody fragments and the subsequent expression, purification and analysis of each particular construct (Arap 2005;Marintcheva 2017). ...
... When phage infection occurs, phage gene expression begins inside the bacterial host, and the inserted peptide/antibody fragment is subsequently displayed on the surface of the phage as a combination product of the relevant genes encoding the coat protein and the cloned sequence (Arap 2005;Aghebati-Maleki et al. 2016). Therefore, if the cloned sequence is randomized, phage display libraries, providing >10 10 variants, can be constructed at the same time and stored for long term as DNA clones, rather than requiring the individual construction of various peptides or antibody fragments and the subsequent expression, purification and analysis of each particular construct (Arap 2005;Marintcheva 2017). The power of phage display technology also arises from its capability to form a physical connection between the displayed molecule (phenotype) and a DNA sequence that encodes the displayed molecule (genotype) (Aghebati-Maleki et al. 2016). ...
... When the expression of a functional peptide/protein is proven, a library of different variants can be developed using mutagenesis methods (Marintcheva 2017). The capability to identify interactive regions of peptides, proteins or antibody fragments without prior knowledge about the type and nature of interaction is a great advantage of the phage display technology, as is the possibility of selecting from a vast number of phage particles in a small sample volume (the titer of phage lysate can be as high as 10 14 virions per ml) due to the small size of phage virions (Arap 2005;Bábíčková et al. 2013). ...
Phage display technology, which is based on the presentation of peptide sequences on the surface of bacteriophage virions, was developed over 30 years ago. Improvements in phage display systems have allowed us to employ this method in numerous fields of biotechnology, as diverse as immunological and biomedical applications, the formation of novel materials and many others. The importance of phage display platforms was recognized by awarding the Nobel Prize in 2018 “for the phage display of peptides and antibodies”. In contrast to many review articles concerning specific applications of phage display systems published in recent years, we present an overview of this technology, including a comparison of various display systems, their advantages and disadvantages, and examples of applications in various fields of science, medicine, and the broad sense of biotechnology. Other peptide display technologies, which employ bacterial, yeast and mammalian cells, as well as eukaryotic viruses and cell-free systems, are also discussed. These powerful methods are still being developed and improved; thus, novel sophisticated tools based on phage display and other peptide display systems are constantly emerging, and new opportunities to solve various scientific, medical and technological problems can be expected to become available in the near future.
... The phage antibody library uses genetic engineering methods to amplify VH and VL genes. After random combination, it is inserted into the phage coat protein gene and fused and expressed on the surface of the phage [20]. Specific single-chain antibodies are obtained through specific panning, which is extensively used for preparing antigen-specific artificial antibodies in biomedicine, environmental pollutants analysis, and food safety detection fields. ...
An immunized mouse phage display scFv library with a capacity of 3.34 × 109 CFU/mL was constructed and used for screening of recombinant anti-ciprofloxacin single-chain antibody for the detection of ciprofloxacin (CIP) in animal-derived food. After four rounds of bio-panning, 25 positives were isolated and identified successfully. The highest positive scFv-22 was expressed in E. coli BL21. Then, its recognition mechanisms were studied using the molecular docking method. The result showed the amino acid residue Val160 was the key residue for the binding of scFv to CIP. Based on the results of virtual mutation, the scFv antibody was evolved by directional mutagenesis of contact amino acid residue Val160 to Ser. After the expression and purification, an indirect competitive enzyme-linked immunosorbent assay (IC-ELISA) based on the parental and mutant scFv was established for CIP, respectively. The IC50 value of the assay established with the ScFv mutant was 1.58 ng/mL, while the parental scFv was 26.23 ng/mL; this result showed highly increased affinity, with up to 16.6-fold improved sensitivity. The mean recovery for CIP ranged from 73.80% to 123.35%, with 10.46% relative standard deviation between the intra-assay and the inter-assay. The RSD values ranged between 1.49% and 9.81%. The results indicate that we obtained a highly sensitive anti-CIP scFv by the phage library construction and directional evolution, and the scFv-based IC-ELISA is suitable for the detection of CIP residue in animal-derived edible tissues.
... The phage display technology is based on the integration of a gene encoding a peptide or a protein fused with the phage coat proteins was first described by George Smith in 1985 [38]. The most broadly used coat proteins for display are the PVIII and PIII proteins; however, other coat proteins likewise been utilized for display. ...
The pandemic of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) was first reported in Wuhan City, China, in 2019. After that, the outbreak has grown into a global pandemic and definite treatment for the disease, termed coronavirus disease 2019 (COVID-19), is currently unavailable. The slow translational progress in the field of research suggests that a large number of studies are urgently required for targeted therapy. In this context, this hypothesis explores the role of bacteriophages on SARS-CoV-2, especially concerning phage therapy (PT). Several studies have confirmed that in addition to their antibacterial abilities, phages also show antiviral properties. It has also been shown that PT is effective for building immunity against viral pathogens by reducing the activation of NF kappa B; additionally, phages produce the antiviral protein phagicin. Phages can also induce antiviral immunity by upregulating expression of defensin 2. Phages may protect eukaryotic cells by competing with viral adsorption and viral penetration of cells, virus mediated cell apoptosis as well as replication. Moreover, by inhibiting activation of NF-κB and ROS production, phages can down regulate excessive inflammatory reactions relevant in clinical course of COVID-19. In this chapter, we hypothesize that the PT may play a therapeutic role in the treatment of COVID-19.
