Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving antigen-antibody binding – specific binding protein...
Patent
1993-04-27
1996-06-11
Ceperley, Mary E.
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving antigen-antibody binding, specific binding protein...
435 793, 436548, 436815, G01N 33577
Patent
active
055254764
DESCRIPTION:
BRIEF SUMMARY
TECHNICAL FIELD
The present invention relates to a method for the determination of a lipophilic (oil-soluble) compound, more particularly relates to a method for the inmunological determination of a lipophilic compound by extracting the lipophilic compound from a sample by an organic solvent, and effecting an antigen-antibody reaction in the presence of an organic solvent, using an organic solvent-resistant antibody against the lipophilic compound.
Furthermore, the present invention relates to a monoclonal antibody specific to diarrheal shellfish poisons, a hybridoma producing the monoclonal antibody, and a method for determining diarrheal shellfish poisons using the monoclonal antibody. The monoclonal antibody according to the present invention is in particular an antibody resistant to an organic solvent.
BACKGROUND ART
In various noxious compounds such as pesticides or toxins, and biologically active important substances such as hormones, there are many compounds which are sparingly soluble in water, but soluble in oil. Hitherto, when the qualitative and quantitative determination of a hydrophobic (water-sparingly-soluble) but lipophilic compound is carried out, the hydrophobic but lipophilic compound to be examined was extracted from a sample by an appropriate organic solvent and purified, and then various instrumental analyses or the like were performed. These procedures were cumbersome and time-consuming in comparison with the case of hydrophilic (water-soluble) compounds which can be directly qualitatively and quantitatively assayed. For example, when assaying okadaic acid which is a toxin contained in marine products, in particular, a diarrheal shellfish poison contained in bivalves such as scallops, the extraction was performed by acetone, ether, ethyl alcohol or methyl alcohol, and the extract was concentrated if necessary, and the assay was carried out by high performance liquid chromatography.
Further, when assaying the above hydrophobic but lipophilic compounds by the immunological method, it was necessary, prior to the determination, to extract the sample possibly containing the substance to be examined by an organic solvent selected in accordance with the extent of the oil solubility of the substance in question. These procedures were cumbersome and time-consuming in comparison with the case of hydrophilic compounds. Further, if there remained the organic solvent used for the extraction, the immunoreaction would be inhibited and thus the reaction would not proceed, or extremely inaccurate results would be obtained in the assay, depending on the concentration of the remaining solvent. To remedy such an inaccurate immunoassay, the organic solvent extract of the lipophilic compound might be diluted with water to lower the concentration of the organic solvent to a concentration where the immunoreaction proceeds accurately (for example, 40% or less). However, even if the immunoreaction could proceed accurately, the solubility of the lipophilic compound per se would be reduced and thus an accurate assay still could not be performed.
Therefore, one of the objects of the present invention is to provide a means enabling swift and specific assay of a hydrophobic but lipophilic compound by immunological means using an antibody.
In the meanwhile, there exist three kinds of diarrheal shellfish poisons; okadaic acid, dinophysistoxin-1, and dinophysistoxin-3 (that is, 7-O-acyl-dinophysistoxin-1). Okadaic acid is a lipophilic compound produced by sponges belonging to Halichondria (Halichondria-okadai and Halichondria-meranodocia), and dinophysistoxin-1 is a lipophilic compound produced by Dinophysis fortii. Further, dinophysistoxin-3 is a lipophilic compound produced by converting dinophysistoxin-1 inside the shellfish. These compounds accumulate in the mesenteron glands of edible bivalves in certain seasons and regions to make the shellfish toxic. The diarrheal shellfish poison is the second most frequent type of food poisoning after blowfish in terms of number of outbreaks, but is the number one in ter
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Fukushi Ken
Hamano Yonekazu
Kabasawa Keigo
Kita Hiroshi
Matsuura Shiro
Ceperley Mary E.
Iatron Laboratories Inc.
Osakafu
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