A White Plaque, Associated with Genomic Deletion, Derived from M13KE-Based Peptide Library Is Enriched in a Target-Unrelated Manner during Phage Display Biopanning Due to Propagation Advantage

Sell, Danna Kamstrup; Sloth, Ane Beth; Bakhshinejad, Babak; Kjær, Andreas

doi:10.3390/ijms23063308

Cited by 4 publications

(3 citation statements)

References 40 publications

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“…While phage display is a powerful technique for identifying epitopes, there are some disadvantages to consider. One potential issue is the risk of false positives or false negatives due to the complexity of the screening process [ 36 ]. This can occur if the phage library is not sufficiently diverse or if the screening conditions are not optimized for the specific antibody or epitope of interest.…”

Section: Resultsmentioning

confidence: 99%

Epitopes and Mimotopes Identification Using Phage Display for Vaccine Development against Infectious Pathogens

Palma¹

2023

Vaccines

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Traditional vaccines use inactivated or weakened forms of pathogens which could have side effects and inadequate immune responses. To overcome these challenges, phage display has emerged as a valuable tool for identifying specific epitopes that could be used in vaccines. This review emphasizes the direct connection between epitope identification and vaccine development, filling a crucial gap in the field. This technique allows vaccines to be engineered to effectively stimulate the immune system by presenting carefully selected epitopes. Phage display involves screening libraries of random peptides or gene/genome fragments using serum samples from infected, convalescent, or vaccinated individuals. This method has been used to identify epitopes from various pathogens including SARS-CoV-2, Mycobacterium tuberculosis, hepatitis viruses, H5N1, HIV-1, Human T-lymphotropic virus 1, Plasmodium falciparum, Trypanosoma cruzi, and Dirofilaria repens. Bacteriophages offer advantages such as being immunogenic carriers, low production costs, and customization options, making them a promising alternative to traditional vaccines. The purpose of this study has been to highlight an approach that encompasses the entire process from epitope identification to vaccine production using a single technique, without requiring additional manipulation. Unlike conventional methods, phage display demonstrates exceptional efficiency and speed, which could provide significant advantages in critical scenarios such as pandemics.

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Section: Resultsmentioning

confidence: 99%

Epitopes and Mimotopes Identification Using Phage Display for Vaccine Development against Infectious Pathogens

Palma¹

2023

Vaccines

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“…To process the FASTQ files, a MATLAB script modified from [ 36 ] was used, as described previously in [ 20 , 37 ]. The script can be found at (accessed on 27 January 2023).…”

Section: Methodsmentioning

confidence: 99%

Depth of Sequencing Plays a Determining Role in the Characterization of Phage Display Peptide Libraries by NGS

Sloth

Bakhshinejad

Stavnsbjerg

et al. 2023

IJMS

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Next-generation sequencing (NGS) has raised a growing interest in phage display research. Sequencing depth is a pivotal parameter for using NGS. In the current study, we made a side-by-side comparison of two NGS platforms with different sequencing depths, denoted as lower-throughput (LTP) and higher-throughput (HTP). The capacity of these platforms for characterization of the composition, quality, and diversity of the unselected Ph.D.TM-12 Phage Display Peptide Library was investigated. Our results indicated that HTP sequencing detects a considerably higher number of unique sequences compared to the LTP platform, thus covering a broader diversity of the library. We found a larger percentage of singletons, a smaller percentage of repeated sequences, and a greater percentage of distinct sequences in the LTP datasets. These parameters suggest a higher library quality, resulting in potentially misleading information when using LTP sequencing for such assessment. Our observations showed that HTP reveals a broader distribution of peptide frequencies, thus revealing increased heterogeneity of the library by the HTP approach and offering a comparatively higher capacity for distinguishing peptides from each other. Our analyses suggested that LTP and HTP datasets show discrepancies in their peptide composition and position-specific distribution of amino acids within the library. Taken together, these findings lead us to the conclusion that a higher sequencing depth can yield more in-depth insights into the composition of the library and provide a more complete picture of the quality and diversity of phage display peptide libraries.

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“…Propagation capacity is determined by an increase in the copy number of phage clones over a given time period during propagation. The higher propagation capacity of individual clones can be attributed to different types of mutations that occur in the phage genome [ 27 , 32 , 33 , 34 ] or the lack of surface display of peptides (wild-type clones (WT) constituting bias in the library construction) [ 35 ]. The presence of phages with faster propagation rates causes a major problem for the library selection, leading to the enrichment of false positive hits—i.e., clones that do not have high binding affinity to the target but dominate the phage pool solely due to increased propagation capacity.…”

Section: Introductionmentioning

confidence: 99%

Propagation Capacity of Phage Display Peptide Libraries Is Affected by the Length and Conformation of Displayed Peptide

et al. 2023

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The larger size and diversity of phage display peptide libraries enhance the probability of finding clinically valuable ligands. A simple way of increasing the throughput of selection is to mix multiple peptide libraries with different characteristics of displayed peptides and use it as biopanning input. In phage display, the peptide is genetically coupled with a biological entity (the phage), and the representation of peptides in the selection system is dependent on the propagation capacity of phages. Little is known about how the characteristics of displayed peptides affect the propagation capacity of the pooled library. In this work, next-generation sequencing (NGS) was used to investigate the amplification capacity of three widely used commercial phage display peptide libraries (Ph.D.™-7, Ph.D.™-12, and Ph.D.™-C7C from New England Biolabs). The three libraries were pooled and subjected to competitive propagation, and the proportion of each library in the pool was quantitated at two time points during propagation. The results of the inter-library competitive propagation assay led to the conclusion that the propagation capacity of phage libraries on a population level is decreased with increasing length and cyclic conformation of displayed peptides. Moreover, the enrichment factor (EF) analysis of the phage population revealed a higher propagation capacity of the Ph.D.TM-7 library. Our findings provide evidence for the contribution of the length and structural conformation of displayed peptides to the unequal propagation rates of phage display libraries and suggest that it is important to take peptide characteristics into account once pooling multiple combinatorial libraries for phage display selection through biopanning.

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A White Plaque, Associated with Genomic Deletion, Derived from M13KE-Based Peptide Library Is Enriched in a Target-Unrelated Manner during Phage Display Biopanning Due to Propagation Advantage

Cited by 4 publications

References 40 publications

Epitopes and Mimotopes Identification Using Phage Display for Vaccine Development against Infectious Pathogens

Epitopes and Mimotopes Identification Using Phage Display for Vaccine Development against Infectious Pathogens

Depth of Sequencing Plays a Determining Role in the Characterization of Phage Display Peptide Libraries by NGS

Propagation Capacity of Phage Display Peptide Libraries Is Affected by the Length and Conformation of Displayed Peptide

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