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 language | English |
|---|---|
| Pages (from-to) | 999-1006 |
| Number of pages | 8 |
| Journal | Biotechnology letters |
| Volume | 28 |
| Issue number | 13 |
| DOIs | |
| Publication status | Published - Jul 1 2006 |
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All Science Journal Classification (ASJC) codes
- Biotechnology
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Activation of a refolded, berberine-specific, single-chain Fv fragment by addition of free berberine. / Kim, Jun Sik; Masaki, Tadashi; Sirikantaramas, Supaart; Shoyama, Yukihiro; Tanaka, Hiroyuki.
In: Biotechnology letters, Vol. 28, No. 13, 01.07.2006, p. 999-1006.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Activation of a refolded, berberine-specific, single-chain Fv fragment by addition of free berberine
AU - Kim, Jun Sik
AU - Masaki, Tadashi
AU - Sirikantaramas, Supaart
AU - Shoyama, Yukihiro
AU - Tanaka, Hiroyuki
PY - 2006/7/1
Y1 - 2006/7/1
N2 - 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.
AB - 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.
UR - http://www.scopus.com/inward/record.url?scp=33745711978&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=33745711978&partnerID=8YFLogxK
U2 - 10.1007/s10529-006-9033-7
DO - 10.1007/s10529-006-9033-7
M3 - Article
C2 - 16786265
AN - SCOPUS:33745711978
VL - 28
SP - 999
EP - 1006
JO - Biotechnology Letters
JF - Biotechnology Letters
SN - 0141-5492
IS - 13
ER -