Bacterial expression and in vitro refolding of a single-chain fv antibody specific for human plasma apolipoprotein B-100

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Title
Bacterial expression and in vitro refolding of a single-chain fv antibody specific for human plasma apolipoprotein B-100
Author(s)
Myung Hoon Lee; Tae In Park; Yong Bok Park; Ju Won Kwak
Bibliographic Citation
Protein Expression and Purification, vol. 25, no. 1, pp. 166-173
Publication Year
2002
Abstract
From the cloned heavy and light chains of a murine monoclonal antibody (mAbB23) which is specific for human apolipoprotein (apo) B-100 of plasma low-density lipoproteins, a vector was designed for expression of a single-chain antibody (scFv) of mAbB23 in Escherichia coli. The expression vector was constructed so that the scFv gene (VL-linker-VH) was expressed under the control of the T7 promoter. The inclusion body of scFv was isolated from E. coli lysate and solubilized in 6 M guanidine-hydrochloride without reducing agents, followed by refolding through slow dilution into refolding buffer. After complete removal of the remaining denaturant by dialysis, the soluble scFv was purified through an apo B-100-coupled affinity column, and an active fraction, which had an antigen-binding activity comparable with that of native Fab, was easily obtained. The expression and in vitro refolding of scFv resulted in production of an active molecule in a yield of 15-20 mg per 1-liter flask cultivation.
Keyword
Apolipoprotein B-100Bacterial expressionIn vitro refoldingmAbB23Single-chain Fv
ISSN
1046-5928
Publisher
Elsevier
DOI
http://dx.doi.org/10.1006/prep.2002.1623
Type
Article
Appears in Collections:
1. Journal Articles > Journal Articles
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