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Protein Eng Des Sel. 2015 Jan;28(1):19-22. doi: 10.1093/protein/gzu049. Epub 2014 Nov 26.

Generation of scFv specific to human VEGFR-3 from the neutralizing mAb BDD073.

Author information

1
Shenzhen Key Laboratory of Genitourinary Tumor, The Second People's Hospital of Shenzhen, The First Affiliated Hospital of Shenzhen University, Shenzhen, P.R. China Faculty of Medicine, School of Biomedical Sciences, The Chinese University of Hong Kong, Shatin, Hong Kong chenhaojob@yahoo.com qihongs@vip.sina.com.
2
Faculty of Medicine, School of Biomedical Sciences, The Chinese University of Hong Kong, Shatin, Hong Kong.
3
National Research Center for Assisted Reproductive Technology and Reproductive Genetics, P.R. China.
4
Department of Immunology, Beijing Institute of Radiation Medicine, Beijing, P.R. China Antibody Engineering Laboratory, Beijing Proteome Research Center, Beijing 100850, P.R. China.
5
Shenzhen Key Laboratory of Genitourinary Tumor, The Second People's Hospital of Shenzhen, The First Affiliated Hospital of Shenzhen University, Shenzhen, P.R. China.
6
Department of Immunology, Beijing Institute of Radiation Medicine, Beijing, P.R. China Antibody Engineering Laboratory, Beijing Proteome Research Center, Beijing 100850, P.R. China chenhaojob@yahoo.com qihongs@vip.sina.com.

Abstract

In our previous study, we have produced a neutralizing mAb of vascular endothelial growth factor receptor 3 (VEGFR-3), specifically BDD073, which could inhibit angiogenesis in the CAM model. However, the clinical application of BDD073 is restricted due to its mouse origin, which might cause human anti-mouse antibody reactions. Herein, we generated functional recombinant single-chain variable fragments (scFv) from mAb BDD073 producing mouse hybridoma cells. The scFv gene containing variable regions of heavy and light chains of BDD073 was cloned into an expression vector with trx tag and expressed in Escherichia coli BL21 (DE3). The recombinant scFv was purified and refolded with Ni-NTA agarose metal affinity column. The bacterially expressed scFv showed moderate potency and specificity to the human VEGFR-3. It may serve as a potential candidate of anti-VEGFR3 treatment for biotechnological and therapeutic applications.

KEYWORDS:

E. coli expression; anti-VEGFR-3; antibody engineering; scFv

PMID:
25428897
DOI:
10.1093/protein/gzu049
[Indexed for MEDLINE]

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