Fluorescence spectroscopy assay means with fluorescent chelate of a lanthanide
First Claim
1. In a method for the determination of a substance by means of fluorescence spectroscopy in which a fluorescence marker is coupled to the molecules of the substance, the fluorescence of the marker having a duration which substantially exceeds the duration of the fluorescence of possible noise sources and in which the substance is excited by means of a short pulse of radiation and the fluorescence thereby generated is detected when the fluorescence from the sources of noise has in principle ceased, the improvement comprising using as said fluorescent marker a 1:
- 1;
1 fluorescent chelate of europium or terbium, β
-diketone or a dihydroxy compound, and an aminopolycarboxylic acid analogue, said chelate having a stability constant above 1010 wherein(a) the europium or terbium is the fluorescence emission source and is chelated to the aminopolycarboxylic acid analogue,(b) the aminopolycarboxylic acid analogue has a functional group for covalent binding to said substance, and(c) said β
-diketone or said dihydroxy compound excites fluorescence of said fluorescence emission source.
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Abstract
An improved method of determining the nature of a substance by fluoroscence spectroscopy wherein a fluorescent marker is coupled to the molecules of the substance comprises the use of a marker having a longer period of fluorescence than those of possible sources of noise and by employing an exciting radiation pulse of short duration so that the fluorescence of the marker is detected after the objectionable sources of fluorescence have ceased; the marker including a fluorescent lanthanide chelate complex.
96 Citations
18 Claims
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1. In a method for the determination of a substance by means of fluorescence spectroscopy in which a fluorescence marker is coupled to the molecules of the substance, the fluorescence of the marker having a duration which substantially exceeds the duration of the fluorescence of possible noise sources and in which the substance is excited by means of a short pulse of radiation and the fluorescence thereby generated is detected when the fluorescence from the sources of noise has in principle ceased, the improvement comprising using as said fluorescent marker a 1:
- 1;
1 fluorescent chelate of europium or terbium, β
-diketone or a dihydroxy compound, and an aminopolycarboxylic acid analogue, said chelate having a stability constant above 1010 wherein(a) the europium or terbium is the fluorescence emission source and is chelated to the aminopolycarboxylic acid analogue, (b) the aminopolycarboxylic acid analogue has a functional group for covalent binding to said substance, and (c) said β
-diketone or said dihydroxy compound excites fluorescence of said fluorescence emission source. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16)
- 1;
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17. In a method for the fluoroimmunoassay of a substance by means of fluorescence spectroscopy in which a fluorescent marker is coupled to the molecules of the substance, the fluorescence of the marker having a duration which substantially exceeds the duration of the fluorescence of possible noise sources and in which the substance is excited by means of a short pulse of radiation and the fluorescence thereby generated is detected when the fluorescence from the sources of noise has in principle ceased, the improvement comprising using as said fluorescent marker a 1:
- 1;
1 fluorescent chelate of europium or terbium, a β
-diketone or a dihydroxy compound, and an aminopolycarboxylic acid analogue, said chelate having a stability constant above 1010 wherein(a) the europium or terbium is the fluorescence emission source and is chelated to the aminopolycarboxylic acid analogue, (b) the aminopolycarboxylic acid has a functional group for covalent binding to said substance, (c) said β
-diketone is selected from the group consisting of benzoylacetone, dibenzoylmethane, thenoyltrifluoroacetone, benzoyltrifluoroacetone, 1- and 2-naphthoyltrifluoroacetone, acetylacetone, trifluoroacetylacetone, and hexafluoroacetylacetone,(d) said dihydroxy compound is selected from the group consisting of sulphosalicylic acid, the disodium salt of 3,5-pyrocatecholdisulfonic acid, 2,3-dihydroxynaphthalene, and 2,3-dihydroxynaphthalene sulfonyl acid derivatives, and (e) the fluorescent 1;
1;
1 structure bound to the substance is formed in the presence of said β
-diketone or said dihydroxy compound. - View Dependent Claims (18)
- 1;
Specification
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- Resources
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Current AssigneeWallac OY A Finnish Joint Stock Company
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Original AssigneeWallac Oy (Revvity, Inc.)
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InventorsHemmila, Ilkka, Soini, Erkki
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Primary Examiner(s)Fagelson, Anna P.
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Application NumberUS06/128,621Time in Patent Office1,075 DaysField of Search424/7, 424/8, 424/12, 424/13, 23/230 B, 250/305, 250/461.2, 435/7, 435/35US Class Current436/546CPC Class CodesG01N 21/6408 with measurement of decay t...G01N 33/582 with fluorescent labelY10S 436/80 Fluorescent dyes, e.g. rhod...
