Construction of a novel phage display antibody library against Fasciola hepatica, and generation of a single-chain variable fragment specific for F. hepatica cathepsin L1

2019 ◽  
Vol 198 ◽  
pp. 87-94 ◽  
Author(s):  
Luke J. Norbury ◽  
Katarzyna Basałaj ◽  
Piotr Bąska ◽  
Alicja Kalinowska ◽  
Anna Zawistowska-Deniziak ◽  
...  
Keyword(s):  
Phage Display ◽  
Fasciola Hepatica ◽  
Single Chain Variable Fragment ◽  
Antibody Library ◽  
Single Chain ◽  
Phage Display Antibody

Generation of a single-chain variable fragment phage display antibody library from naïve mice panned against Fasciola hepatica antigens

2019 ◽  
Vol 205 ◽  
pp. 107737
Author(s):  
Luke J. Norbury ◽  
Katarzyna Basałaj ◽  
Piotr Bąska ◽  
Anna Zawistowska-Deniziak ◽  
Alicja Kalinowska ◽  
...  
Keyword(s):  
Phage Display ◽  
Fasciola Hepatica ◽  
Single Chain Variable Fragment ◽  
Antibody Library ◽  
Single Chain ◽  
Phage Display Antibody

Screening of Single-Chain Variable Fragments against TSP50 from a Phage Display Antibody Library and Their Expression as Soluble Proteins

2006 ◽  
Vol 11 (5) ◽  
pp. 546-552 ◽  
Author(s):  
Jingyan Wei ◽  
Yang Liu ◽  
Songchuan Yang ◽  
Junjie Xu ◽  
Hangtian Kong ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Phage Display ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sandwich Elisa ◽  
Sodium Dodecyl ◽  
Enzyme Linked Immunosorbent Assay ◽  
Antibody Library ◽  
Single Chain ◽  
Single Chain Variable Fragments ◽  
Phage Display Antibody

A novel gene, testes-specific protease 50 ( TSP50), is abnormally activated and differentially expressed in most patients with breast cancer, suggesting it as a novel biomarker for this disease. The possibility that TSP50 may be an oncogene is presently under investigation. In this study, the single-chain variable fragments (scFvs) against TSP50 were panned from a phage display antibody library using TSP50-specific peptide, pep-50, as a target antigen. After 4 rounds of panning, 3 clones (A1, A11, and C8) from the library were verified to show strong binding affinities for TSP50 by enzyme-linked immunosorbent assay (ELISA) and to contain the variable region genes of the light and heavy chains of scFv antibodies but different complementary determining regions by sequencing. The genes of scFv-A1 and scFv-A11 were cloned into expression vector pPELB and successfully expressed as a soluble protein in Escherichia coli Rosetta. The yields of expressions were about 4.0 to 5.0 mg of protein from 1 L of culture. The expressed proteins were purified by a 2-step procedure consisting of ion-exchange chromatography, followed by immobilized metal affinity chromatography. The purified proteins were shown a single band at the position of 31 KDa on sodium dodecyl sulfate–polyacrylamide gel electrophoresis. Sandwich ELISA demonstrated that the expressed scFv proteins were able to specifically react with pep-50, laying a foundation for the investigation of the function of TSP50 in the development and treatment of breast cancer.

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