Improved Soluble ScFv ELISA Screening Approach for Antibody Discovery Using Phage Display Technology.
SLAS Discov
; 22(8): 1026-1034, 2017 09.
Article
en En
| MEDLINE
| ID: mdl-28346811
ABSTRACT
Phage display technology (PDT) is a powerful tool for the isolation of recombinant antibody (Ab) fragments. Using PDT, target molecule-specific phage-Ab clones are enriched through the "biopanning" process. The individual specific binders are screened by the monoclonal scFv enzyme-linked immunosorbent assay (ELISA) that may associate with inevitable false-negative results. Thus, in this study, three strategies were investigated for optimization of the scFvs screening using Tomlinson I and J libraries, including (1) optimizing the expression of functional scFvs, (2) improving the sensitivity of ELISA, and (3) preparing different samples containing scFvs. The expression of all scFv Abs was significantly enhanced when scFv clones were cultivated in the terrific broth (TB) medium at the optimum temperature of 30 °C. The protein A-conjugated with horseradish peroxidase (HRP) was found to be a well-suited reagent for the detection of Ag-bound scFvs in comparison with either anti-c-myc Ab or the mixing procedure. Based on our findings, it seems there is no universal media supplement for an improved expression of all scFvs derived from both Tomlinson I and J libraries. We thus propose that expression of scFv fragments in a microplate scale is largely dependent on a variety of parameters, in particular the scFv clones and relevant sequences.
Palabras clave
Texto completo:
1
Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Ensayo de Inmunoadsorción Enzimática
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Biblioteca de Péptidos
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Descubrimiento de Drogas
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Anticuerpos de Cadena Única
Tipo de estudio:
Diagnostic_studies
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Screening_studies
Límite:
Humans
Idioma:
En
Revista:
SLAS Discov
Año:
2017
Tipo del documento:
Article
País de afiliación:
Irán
Pais de publicación:
EEUU
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ESTADOS UNIDOS
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ESTADOS UNIDOS DA AMERICA
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EUA
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UNITED STATES
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UNITED STATES OF AMERICA
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US
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USA