scholarly journals Oxford nanopore sequencing enables rapid discovery of single-domain antibodies from phage display libraries

BioTechniques ◽  
2018 ◽  
Vol 65 (6) ◽  
pp. 351-356 ◽  
Author(s):  
Michael J Lowden ◽  
Kevin A Henry
Keyword(s):  
Phage Display ◽  
Single Domain ◽  
Nanopore Sequencing ◽  
Phage Display Libraries ◽  
Oxford Nanopore ◽  
Single Domain Antibodies

scholarly journals Isolation of rabbit single domain antibodies to B7-H3 via protein immunization and phage display

2020 ◽  
Vol 3 (1) ◽  
pp. 10-17
Author(s):  
Ruonan Feng ◽  
Ruixue Wang ◽  
Jessica Hong ◽  
Christopher M Dower ◽  
Brad St Croix ◽  
...  
Keyword(s):  
Tumor Cell ◽  
Phage Display ◽  
Cell Lines ◽  
Single Domain ◽  
Tumor Cell Lines ◽  
Positive Tumor Cell ◽  
Drug Candidates ◽  
Potential Applications ◽  
Single Domain Antibodies ◽  
Protein Immunization

Abstract Single domain antibodies have certain advantages including their small size, high stability and excellent tissue penetration, making them attractive drug candidates. Rabbit antibodies can recognize diverse epitopes, including those that are poorly immunogenic in mice and humans. In the present study, we established a method to isolate rabbit VH single domain antibodies for potential cancer therapy. We immunized rabbits with recombinant human B7-H3 (CD276) protein, made a phage-displayed rabbit VH single domain library with a diversity of 7 × 109, and isolated two binders (A1 and B1; also called RFA1 and RFB1) from phage panning. Both rabbit VH single domains exhibited antigen-dependent binding to B7-H3-positive tumor cell lines but not B7-H3 knockout tumor cell lines. Our study shows that protein immunization followed by phage display screening can be used to isolate rabbit single domain antibodies. The two single domain antibodies reported here may have potential applications in cancer immunotherapy.

scholarly journals Identification of Novel Single-Domain Antibodies against FGF7 Using Phage Display Technology

2017 ◽  
Vol 23 (2) ◽  
pp. 193-201
Author(s):  
Behzad Jafari ◽  
Maryam Hamzeh-Mivehroud ◽  
Ali A. Moosavi-Movahedi ◽  
Siavoush Dastmalchi
Keyword(s):  
Growth Factor ◽  
Phage Display ◽  
Fibroblast Growth Factor ◽  
Single Domain ◽  
Enzyme Linked Immunosorbent Assay ◽  
Fibroblast Growth ◽  
Phage Display Technology ◽  
Display Technology ◽  
Single Domain Antibodies

Fibroblast growth factor 7 (FGF7) is a member of the fibroblast growth factor (FGF) family of proteins. FGF7 is of stromal origin and produces a paracrine effect on epithelial cells. In the current investigation, we aimed to identify new single-domain antibodies (sdAbs) against FGF7 using phage display technology. The vector harboring the codon-optimized DNA sequence for FGF7 protein was transformed into Escherichia coli BL21 (DE3) pLysS, and then the protein was expressed at the optimized condition. Enzyme-linked immunosorbent assay, circular dichroism spectropolarimetry, and in vitro scratch assay experiments were used to confirm the proper folding and functionality of the purified FGF7 protein. The purity of the produced FGF7 was 92%, with production yield of 3.5 mg/L of culture. Panning against the purified FGF7 was performed, and the identified single-domain antibodies showed significant affinity. Further investigation on one of the selected sdAb displaying phage clones showed concentration-dependent binding to FGF7. The selected sdAb can be used for developing novel tumor-suppressing agents where inhibition of FGF7 is required.

Computer-guided library generation applied to the optimization of single-domain antibodies

2019 ◽  
Vol 32 (9) ◽  
pp. 423-431
Author(s):  
Hiroki Akiba ◽  
Hiroko Tamura ◽  
Jose M M Caaveiro ◽  
Kouhei Tsumoto
Keyword(s):  
Phage Display ◽  
In Silico ◽  
Single Domain ◽  
Alanine Scanning ◽  
Alanine Residue ◽  
The Core ◽  
Computational Alanine Scanning ◽  
Single Domain Antibodies ◽  
Antigen Interaction ◽  
Interaction Surface

Abstract Computer-guided library generation is a plausible strategy to optimize antibodies. Herein, we report the improvement of the affinity of a single-domain camelid antibody for its antigen using such approach. We first conducted experimental and computational alanine scanning to describe the precise energetic profile of the antibody–antigen interaction surface. Based on this characterization, we hypothesized that in-silico mutagenesis could be employed to guide the development of a small library for phage display with the goal of improving the affinity of an antibody for its antigen. Optimized antibody mutants were identified after three rounds of selection, in which an alanine residue at the core of the antibody–antigen interface was substituted by residues with large side-chains, generating diverse kinetic responses, and resulting in greater affinity (>10-fold) for the antigen.

scholarly journals A Novel Affinity Tag, ABTAG, and Its Application to the Affinity Screening of Single-Domain Antibodies Selected by Phage Display

2017 ◽  
Vol 8 ◽  
Author(s):  
Greg Hussack ◽  
Toya Nath Baral ◽  
Jason Baardsnes ◽  
Henk van Faassen ◽  
Shalini Raphael ◽  
...  
Keyword(s):  
Phage Display ◽  
Single Domain ◽  
Affinity Tag ◽  
Single Domain Antibodies

Retooling phage display with electrohydrodynamic nanomixing and nanopore sequencing

Lab on a Chip ◽  
2019 ◽  
Vol 19 (24) ◽  
pp. 4083-4092 ◽  
Author(s):  
Lyndon J. Raftery ◽  
Christopher B. Howard ◽  
Yadveer S. Grewal ◽  
Ramanathan Vaidyanathan ◽  
Martina L. Jones ◽  
...  
Keyword(s):  
Phage Display ◽  
High Throughput ◽  
High Throughput Screening ◽  
Nanopore Sequencing ◽  
Phage Display Libraries ◽  
Target Binding

High throughput screening of phage display libraries for target binding molecules using electrohydrodynamic nanomixing and nanopore sequencing.

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