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Bacterial Inner-membrane Display for Screening a Library of Antibody Fragments
Published on: October 15, 2016
Screening of high-affinity scFvs from a ribosome displayed library using BIAcore biosensor
Qing Yuan1, Zhongkang Wang, Siji Nian
1Genetic Engineering Research Centre, College of Bioengineering, Chongqing University, People's Republic of China.
Applied Biochemistry and Biotechnology
|June 25, 2008
Summary
Researchers developed a BIAcore biosensor method to screen high-affinity single-chain variable fragment (scFv) antibodies against Xanthomonas axonopodis pv. citri (Xac) lipopolysaccharides (LPS). Three scFvs demonstrated high affinity and specificity for Xac LPS, validating the screening approach.
Area of Science:
- Biotechnology
- Immunology
- Molecular Biology
Background:
- Xanthomonas axonopodis pv. citri (Xac) causes citrus canker, necessitating effective diagnostic tools.
- Ribosome display and BIAcore biosensor technology offer powerful platforms for antibody fragment screening.
Purpose of the Study:
- To develop and validate an experimental protocol for screening high-affinity single-chain Fv antibody fragments (scFvs) against Xac.
- To identify specific scFvs targeting lipopolysaccharides (LPS) of Xac using a BIAcore biosensor.
Main Methods:
- Utilized ribosome display library and BIAcore biosensor for screening scFvs against immobilized Xac LPS.
- Selected scFvs based on dissociation rate constants (k (d)) and affinity constants.
- Expressed and purified selected scFvs in Escherichia coli for further validation.
Main Results:
- Identified three high-affinity scFvs (A1, B2, C5) with dissociation constants in the nanomolar range (10⁻¹⁰ to 10⁻¹¹ M).
- ScFv B2 exhibited high specificity for Xac LPS, showing no cross-reactivity with other antigens.
- BIAcore screening assay results were validated through purified scFv characterization.
Conclusions:
- The developed BIAcore screening protocol accurately identifies high-affinity and specific scFvs against Xac LPS.
- The identified scFvs, particularly B2, hold potential for developing diagnostic tools for Xac detection.
