NDLI: Generation and characterization of a human single-chain fragment variable (scFv) antibody against cytosine deaminase from Yeast

Content Provider	Springer Nature : BioMed Central
Author	Mallano, Alessandra Zamboni, Silvia Carpinelli, Giulia Santoro, Filippo Flego, Michela Ascione, Alessandro Gellini, Mara Tombesi, Marina Podo, Franca Cianfriglia, Maurizio
Abstract	Background The ability of cytosine deaminase (CD) to convert the antifungal agent 5-fluorocytosine (5-FC) into one of the most potent and largely used anticancer compound such as 5-fluorouracil (5-FU) raised considerable interest in this enzyme to model gene or antibody – directed enzyme-prodrug therapy (GDEPT/ADEPT) aiming to improve the therapeutic ratio (benefit versus toxic side-effects) of cancer chemotherapy. The selection and characterization of a human monoclonal antibody in single chain fragment (scFv) format represents a powerful reagent to allow in in vitro and in vivo detection of CD expression in GDEPT/ADEPT studies. Results An enzymatic active recombinant CD from yeast (yCD) was expressed in E. coli system and used as antigen for biopanning approach of the large semi-synthetic ETH-2 antibody phage library. Several scFvs were isolated and specificity towards yCD was confirmed by Western blot and ELISA. Further, biochemical and functional investigations demonstrated that the binding of specific scFv with yCD did not interfere with the activity of the enzyme in converting 5-FC into 5-FU. Conclusion The construction of libraries of recombinant antibody fragments that are displayed on the surface of filamentous phage, and the selection of phage antibodies against target antigens, have become an important biotechnological tool in generating new monoclonal antibodies for research and clinical applications. The scFvH5 generated by this method is the first human antibody which is able to detect yCD in routinary laboratory techniques without interfering with its enzymatic function.
Related Links	https://bmcbiotechnol.biomedcentral.com/counter/pdf/10.1186/1472-6750-8-68.pdf
Ending Page	10
Page Count	10
Starting Page	1
File Format	HTM / HTML
ISSN	14726750
DOI	10.1186/1472-6750-8-68
Journal	BMC Biotechnology
Issue Number	1
Volume Number	8
Language	English
Publisher	BioMed Central
Publisher Date	2008-09-10
Access Restriction	Open
Subject Keyword	Applied Microbiology Biotechnology Biochemical Engineering Genetic Engineering Plant Breeding Biomedical Engineering LoVo Cell Cytosine Deaminase scFv Antibody Interpulse Delay Soluble scFv Plant Breeding/Biotechnology Biomedical Engineering/Biotechnology
Content Type	Text
Resource Type	Article
Subject	Biotechnology
Journal Impact Factor	3.5/2023
5-Year Journal Impact Factor	3.2/2023

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