Activation of a refolded, berberine-specific, single-chain Fv fragment by addition of free berberine

Jun Sik Kim, Tadashi Masaki, Supaart Sirikantaramas, Yukihiro Shoyama, Hiroyuki Tanaka

Research output: Contribution to journalArticle

3 Citations (Scopus)

Abstract

A single-chain variable fragment (scFv) specific for berberine was produced in Escherichia coli. The anti-berberine scFv gene was cloned from hybridoma 1D5-3B-7 producing the monoclonal antibody. The variable regions of the heavy (VH) and light chain (VL) genes were connected with a flexible linker using an assembly PCR. The VH-linker-VL gene was inserted into a plasmid, pET28a (+), then overexpressed in E. coli BL21 (DE3). The active of the scFv by refolding based on stepwise dialysis methods and an artificial chaperone was determined by direct and competitive enzyme-linked immunosorbent assay (ELISA). The results of direct ELISA showed that the anti-berberine scFv retained specific binding activity to berberine. In competitive ELISA, however, activity was increased depending on the concentration of berberine.

Original languageEnglish
Pages (from-to)999-1006
Number of pages8
JournalBiotechnology letters
Volume28
Issue number13
DOIs
Publication statusPublished - Jul 1 2006

All Science Journal Classification (ASJC) codes

  • Biotechnology

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