Abstract
A novel automated flow immunoassay system is developed for the detection of wheat protein as a food allergen. A polyclonal immunoglobulin G (IgG) antibody was isolated from serum of a rat with a wheat allergy, then reductively treated using dithiothreitol and covalently coupled to alkaline phosphatase. This conjugate was used in a non-competitive binding assay. The automated system was equipped with an immunoreaction column, an anion-exchange column, a luminescent reaction column and a photodetector. Antibody-antigen complexes were separated from their free conjugate on the basis of a difference in isoelectric point (pI) by an anion-exchange column. This simple technique permits the assay of wheat-protein allergen in 25min with a reliable detection limit of 5μg/ml. The anion-exchange column was regenerated by occasional elution with N-methylpiperazine buffer (pH 5.0) containing 0.5M NaCl, to remove free conjugate. Free conjugate recovered in this manner could be reused up to four times without significant decrease in sensitivity of the immunoassay. Copyright (C) 1998 Elsevier Science B.V.
| Original language | English |
|---|---|
| Pages (from-to) | 207-214 |
| Number of pages | 8 |
| Journal | Analytica Chimica Acta |
| Volume | 370 |
| Issue number | 2-3 |
| DOIs | |
| Publication status | Published - 1998 Sep 7 |
| Externally published | Yes |
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Keywords
- Alkaline phosphatase
- Anion-exchange resin
- Automated flow immuno assay
- Wheat allergen
ASJC Scopus subject areas
- Biochemistry
- Analytical Chemistry
- Spectroscopy
- Environmental Chemistry
Cite this
Automated flow immunoassay for detection of food allergen using anion-exchange resin and alkaline phosphatase conjugated immunoglobulin G. / Lim, Tae Kyu; Nakamura, Noriyuki; Matsunaga, Tadashi.
In: Analytica Chimica Acta, Vol. 370, No. 2-3, 07.09.1998, p. 207-214.Research output: Contribution to journal › Article
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TY - JOUR
T1 - Automated flow immunoassay for detection of food allergen using anion-exchange resin and alkaline phosphatase conjugated immunoglobulin G
AU - Lim, Tae Kyu
AU - Nakamura, Noriyuki
AU - Matsunaga, Tadashi
PY - 1998/9/7
Y1 - 1998/9/7
N2 - A novel automated flow immunoassay system is developed for the detection of wheat protein as a food allergen. A polyclonal immunoglobulin G (IgG) antibody was isolated from serum of a rat with a wheat allergy, then reductively treated using dithiothreitol and covalently coupled to alkaline phosphatase. This conjugate was used in a non-competitive binding assay. The automated system was equipped with an immunoreaction column, an anion-exchange column, a luminescent reaction column and a photodetector. Antibody-antigen complexes were separated from their free conjugate on the basis of a difference in isoelectric point (pI) by an anion-exchange column. This simple technique permits the assay of wheat-protein allergen in 25min with a reliable detection limit of 5μg/ml. The anion-exchange column was regenerated by occasional elution with N-methylpiperazine buffer (pH 5.0) containing 0.5M NaCl, to remove free conjugate. Free conjugate recovered in this manner could be reused up to four times without significant decrease in sensitivity of the immunoassay. Copyright (C) 1998 Elsevier Science B.V.
AB - A novel automated flow immunoassay system is developed for the detection of wheat protein as a food allergen. A polyclonal immunoglobulin G (IgG) antibody was isolated from serum of a rat with a wheat allergy, then reductively treated using dithiothreitol and covalently coupled to alkaline phosphatase. This conjugate was used in a non-competitive binding assay. The automated system was equipped with an immunoreaction column, an anion-exchange column, a luminescent reaction column and a photodetector. Antibody-antigen complexes were separated from their free conjugate on the basis of a difference in isoelectric point (pI) by an anion-exchange column. This simple technique permits the assay of wheat-protein allergen in 25min with a reliable detection limit of 5μg/ml. The anion-exchange column was regenerated by occasional elution with N-methylpiperazine buffer (pH 5.0) containing 0.5M NaCl, to remove free conjugate. Free conjugate recovered in this manner could be reused up to four times without significant decrease in sensitivity of the immunoassay. Copyright (C) 1998 Elsevier Science B.V.
KW - Alkaline phosphatase
KW - Anion-exchange resin
KW - Automated flow immuno assay
KW - Wheat allergen
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U2 - 10.1016/S0003-2670(98)00288-8
DO - 10.1016/S0003-2670(98)00288-8
M3 - Article
AN - SCOPUS:0031878116
VL - 370
SP - 207
EP - 214
JO - Analytica Chimica Acta
JF - Analytica Chimica Acta
SN - 0003-2670
IS - 2-3
ER -