Targeting Cytokines: Production and Characterization of Anti-TNF-α scFvs by Phage Display Technology

Abstract Background: Different molecular forms of prostate-specific antigen (PSA) have been used to differentiate between benign prostatic hyperplasia and prostate cancer. Detecting PSA bound to endogenous inhibitors such as α1-antichymotrypsin (ACT) and α2-macroglobulin (α2M) is often difficult because of epitope masking or sensitivity problems. Here we report the characterization of four novel mouse monoclonal antibodies (mabs) obtained by immunization with PSA-α2M complexes. Their ability to detect free PSA and PSA-inhibitor complexes was shown, and their epitopes were analyzed by phage display technology. Methods: The properties of the mabs were studied by competition and sandwich assays and by Western blotting. Epitope mapping was performed by screening of a phage display peptide library. Results: All four mabs recognized free PSA, PSA-ACT, and PSA-α2M complexes, but to various degrees. With different combinations of mabs in competition experiments, antibodies were identified that enhance binding of other mabs to PSA, forming the molecular basis of a very sensitive assay for the detection of PSA and PSA-ACT complexes. Mabs with highest reactivity for PSA-α2M were selected to establish an immunoassay for that complex. Western blot analysis revealed that all mabs recognized conformational epitopes of PSA. These findings were supported by phage display results demonstrating mimotopes in the PSA molecule. Conclusion: The results presented here could aid in the further development of clinically relevant assays for PSA and PSA-α2M complexes.

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Screening and characterization of activin A inhibitory peptides by phage display technology

10.1183/13993003.congress-2018.pa2963 ◽

2018 ◽

Author(s):

Victor Alexandre Félix Bastos ◽

Aline Gomes Souza ◽

Patrícia Tieme Fujimura ◽

Robinson Sabino Silva ◽

Thulio Marquez Cunha ◽

...

Keyword(s):

Phage Display ◽

Activin A ◽

Phage Display Technology ◽

Inhibitory Peptides ◽

Display Technology

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Development and Application of Computational Methods in Phage Display Technology

Current Medicinal Chemistry ◽

10.2174/0929867325666180629123117 ◽

2020 ◽

Vol 26 (42) ◽

pp. 7672-7693 ◽

Cited By ~ 3

Author(s):

Bifang He ◽

Anthony Mackitz Dzisoo ◽

Ratmir Derda ◽

Jian Huang

Keyword(s):

Phage Display ◽

Computational Methods ◽

Peptide Ligands ◽

Next Generation ◽

Phage Display Technology ◽

Next Generation Sequencing Technology ◽

Screening Experiments ◽

Display Technology ◽

Comprehensive Search ◽

Generation Sequencing

Background: Phage display is a powerful and versatile technology for the identification of peptide ligands binding to multiple targets, which has been successfully employed in various fields, such as diagnostics and therapeutics, drug-delivery and material science. The integration of next generation sequencing technology with phage display makes this methodology more productive. With the widespread use of this technique and the fast accumulation of phage display data, databases for these data and computational methods have become an indispensable part in this community. This review aims to summarize and discuss recent progress in the development and application of computational methods in the field of phage display. Methods: We undertook a comprehensive search of bioinformatics resources and computational methods for phage display data via Google Scholar and PubMed. The methods and tools were further divided into different categories according to their uses. Results: We described seven special or relevant databases for phage display data, which provided an evidence-based source for phage display researchers to clean their biopanning results. These databases can identify and report possible target-unrelated peptides (TUPs), thereby excluding false-positive data from peptides obtained from phage display screening experiments. More than 20 computational methods for analyzing biopanning data were also reviewed. These methods were classified into computational methods for reporting TUPs, for predicting epitopes and for analyzing next generation phage display data. Conclusion: The current bioinformatics archives, methods and tools reviewed here have benefitted the biopanning community. To develop better or new computational tools, some promising directions are also discussed.

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