Abstract
In the development of immunoassay technique, the design of hapten containing a functional group suitable for protein conjugate is the key step for the preparation of antibodies against small molecules. Coptisine (MW 320), a bioactive constituent of Berberis and Coptis species, is small as an immunogen. In addition, coptisine has no reactive group in molecule for conjugating with a protein. To overcome this problem, 9-O-carboxymethyl-berberrubine was designed and conjugated with carrier protein. In order to confirm its immunogenicity, the ratio of hapten in the conjugate was determined by matrix-assisted laser desorption/ ionization time of flight mass spectrometry (MALDI-TOF MS). After immunization, hybridomas secreting antibodies against coptisine were produced by fusing splenocytes with mouse myeloma cell line, P3-X63-Ag8-653. Among hybridomas, the clone 2A1 secreting anti-coptisine monoclonal antibody (MAb) 2A1-9E-1 was obtained through the limited dilution method. The MAb-based enzyme-linked immunosorbent assay (ELISA) against coptisine was developed and characterized. The linear range of the assay in this ELISA method was extended from 1.56 to 25 μg ml -1 possessing the detection limit of 1.56 μg ml -1 . The established ELISA using MAb 2A1-9E-1 was applied for the survey of isoquinoline alkaloids in various medicinal plants.
| Original language | English |
|---|---|
| Pages (from-to) | 115-123 |
| Number of pages | 9 |
| Journal | Cytotechnology |
| Volume | 44 |
| Issue number | 3 |
| DOIs | |
| Publication status | Published - Dec 1 2004 |
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All Science Journal Classification (ASJC) codes
- Biotechnology
- Bioengineering
- Biomedical Engineering
- Clinical Biochemistry
- Cell Biology
Cite this
Development of monoclonal antibody against isoquinoline alkaloid coptisine and its application for the screening of medicinal plants. / Kim, Jun Sik; Tanaka, Hiroyuki; Yuan, Chun Su; Shoyama, Yukihiro.
In: Cytotechnology, Vol. 44, No. 3, 01.12.2004, p. 115-123.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Development of monoclonal antibody against isoquinoline alkaloid coptisine and its application for the screening of medicinal plants
AU - Kim, Jun Sik
AU - Tanaka, Hiroyuki
AU - Yuan, Chun Su
AU - Shoyama, Yukihiro
PY - 2004/12/1
Y1 - 2004/12/1
N2 - In the development of immunoassay technique, the design of hapten containing a functional group suitable for protein conjugate is the key step for the preparation of antibodies against small molecules. Coptisine (MW 320), a bioactive constituent of Berberis and Coptis species, is small as an immunogen. In addition, coptisine has no reactive group in molecule for conjugating with a protein. To overcome this problem, 9-O-carboxymethyl-berberrubine was designed and conjugated with carrier protein. In order to confirm its immunogenicity, the ratio of hapten in the conjugate was determined by matrix-assisted laser desorption/ ionization time of flight mass spectrometry (MALDI-TOF MS). After immunization, hybridomas secreting antibodies against coptisine were produced by fusing splenocytes with mouse myeloma cell line, P3-X63-Ag8-653. Among hybridomas, the clone 2A1 secreting anti-coptisine monoclonal antibody (MAb) 2A1-9E-1 was obtained through the limited dilution method. The MAb-based enzyme-linked immunosorbent assay (ELISA) against coptisine was developed and characterized. The linear range of the assay in this ELISA method was extended from 1.56 to 25 μg ml -1 possessing the detection limit of 1.56 μg ml -1 . The established ELISA using MAb 2A1-9E-1 was applied for the survey of isoquinoline alkaloids in various medicinal plants.
AB - In the development of immunoassay technique, the design of hapten containing a functional group suitable for protein conjugate is the key step for the preparation of antibodies against small molecules. Coptisine (MW 320), a bioactive constituent of Berberis and Coptis species, is small as an immunogen. In addition, coptisine has no reactive group in molecule for conjugating with a protein. To overcome this problem, 9-O-carboxymethyl-berberrubine was designed and conjugated with carrier protein. In order to confirm its immunogenicity, the ratio of hapten in the conjugate was determined by matrix-assisted laser desorption/ ionization time of flight mass spectrometry (MALDI-TOF MS). After immunization, hybridomas secreting antibodies against coptisine were produced by fusing splenocytes with mouse myeloma cell line, P3-X63-Ag8-653. Among hybridomas, the clone 2A1 secreting anti-coptisine monoclonal antibody (MAb) 2A1-9E-1 was obtained through the limited dilution method. The MAb-based enzyme-linked immunosorbent assay (ELISA) against coptisine was developed and characterized. The linear range of the assay in this ELISA method was extended from 1.56 to 25 μg ml -1 possessing the detection limit of 1.56 μg ml -1 . The established ELISA using MAb 2A1-9E-1 was applied for the survey of isoquinoline alkaloids in various medicinal plants.
UR - http://www.scopus.com/inward/record.url?scp=12144252780&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=12144252780&partnerID=8YFLogxK
U2 - 10.1007/s10616-004-1179-3
DO - 10.1007/s10616-004-1179-3
M3 - Article
VL - 44
SP - 115
EP - 123
JO - Cytotechnology
JF - Cytotechnology
SN - 0920-9069
IS - 3
ER -