Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving oxidoreductase
Patent
1991-02-05
1993-04-27
Wityshyn, Michael G.
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving oxidoreductase
435 6, 435 25, 435 772, 435 5, 25230117, 536 181, 514212, 540524, C12Q 168, C12Q 128, G01N 5300, G01N 2152
Patent
active
052061493
DESCRIPTION:
BRIEF SUMMARY
BACKGROUND OF THE INVENTION
1. Technical Field
This invention relates to a method of high sensitivity analysis through chemiluminescence measurement in use of enzymatic reactions, antigen-antibody reactions and nucleic acid hybridization in the areas of clinical laboratory testing, food inspection, environmental analysis, inspection of animals and plants, and manufacturing process control checking.
2. Background Art
The luminescent reaction using oxidation of luminol, isoluminol or a derivative thereof [abbreviated as chemiluminescent DPD (2,3-dihydro-1,4-phthalazinedione) in the following] by a peroxidase is used for immunoassay, analysis of elastase, analysis of glucose and analysis of oxidants. It is known that for improving the luminescent intensity of said luminescent reaction, it is effective to add an enhancer, such as those shown below, to the reaction system. Publication No. SHO 59-171839)
However, such enhancers have the following difficulties.
The enhancer (1) is generally of lower luminescence intensity and smaller signal-to-background ratio.
The typical enhancer of the class (2) is p-iodophenol. Its luminescent signal is high, but the background is also high, therefore the signal-to-background ratio is relatively low.
In the case of the typical enhancer of the class (3) or N,N,N',N'-tetramethylbenzidine, the rise to the luminescent peak is slow, and much time is required for measurement.
For the reaction mechanism of the enhancer effect, there are reports proposing the requirement of efficient formation of luminol semiquinone radical [Thorpe, G. H. C. and Kricka, L. J., "Bioluminescence and Chemiluminescence: New Perspectives," Scholmerich, J., Andreesn, R., Kapp, A., Ernst, M. and Woods, W. G. (Eds), John Wiley, Chichester, pp. 199-208 (1987)] and the requirement of efficient formation of phenoxy radical [Hodgson, M. and Jones, P., Journal of Bioluminescence and Chemiluminescence, Vol. 3, pp. 21-25 (1989)].
However, the phenol derivatives include a number of compounds which do not show an enhancer effect, and it is difficult to choose an effective enhancer based upon the foregoing theories.
Also, for detection or quantification of a gene in a virulent microorganism, a prostaglandin or any other physiologically active substance, luteinizing hormone (LH) and other anterior pituitary hormones, and cytokines such as interleukin in blood, it is required to make a determination based on very small amount in body fluid. Thus, a high sensitivity detection system was called for, and for improvement of the detection sensitivity, an enhancer of higher efficiency has been desired.
SUMMARY OF THE INVENTION
An object of the present invention is to provide a method of high sensitivity luminescent analysis which is characteristically carried out in the presence of a novel enhancer.
Another object of the present invention is to provide a novel oxazole derivative which is effective as an enhancer and allows the luminescent analysis to be carried out with an enhancer of higher efficiency than the conventional enhancers used.
In detecting or determining a substance in use of the chemiluminescence produced through reaction of (a) a peroxidase or a derivative thereof, (b) an oxidant and (c) luminol or isoluminol, or a derivative thereof, the present invention relates to a method of high sensitivity luminescent analysis characterized in that the luminescence inducing reaction is carried out in the presence of at least one compound chosen from the group comprised of 2-hydroxy-9-fluorenone, the compound expressed by the following formula ##STR3## and oxazole derivatives expressed by the formula ##STR4## in the formula, R.sub.1 representing hydrogen, C.sub.n H.sub.2n+1 (here, n representing a positive integer of 1 to 4), XC.sub.n H.sub.2n (here, X representing F, Cl, Br or I; n being as defined in the foregoing), C.sub.n H.sub.2n+1 CO.sub.2 (n being as defined in the foregoing), phenyl group, naphthyl group, C.sub.n H.sub.2n+1 C.sub.6 H.sub.4 (n being as defined in the foregoing), YC.sub.6 H.sub.4 (Y representing
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Hosaka Shuntaro
Makino Tetsuya
Oyama Yoshihiro
Leary Louise N.
Toray Industries Inc.
Wityshyn Michael G.
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