Energy transfer hybridization assay using intercalators and lanthanide metals
First Claim
1. A nucleic acid hybridization composition comprising:
- an oligo- or polynucleotide having directly or indirectly bound thereto at least one lanthanide metal or at least one fluorophore, each acting as either an energy donor or an energy acceptor;
a solid matrix having at least one surface to which is attached a first intercalator capable of capturing a double-stranded nucleic acid;
a second intercalator, that may or may not comprise at least one fluorophore, said second intercalator or said fluorophore, each acting as either an energy donor or an energy acceptor wherein upon hybridization of said oligo- or polynucleotide to a complementary oligo- or polynucleotide, said energy donor and said energy acceptor are within proximate distance of each other such that energy from said energy donor is absorbed by said energy acceptor.
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Abstract
Disclosed is a nucleic acid hybridization assay composition for detecting the presence of absence of a target oligo- or polynucleotide in a sample. The composition comprises: a solid matrix having at least one surface which is substituted with a first intercalator capable of binding dsDNA dsRNA, or DAN-RNA hybrids; a second intercalator, which may or may not comprise at least one fluorophore, said intercalator or said fluorophore each acting as either an energy donor or an energy acceptor; and an oligo- or polynucleotide probe which is specifically hybridizable with the target oligo- or polynucleotide and has directly or indirectly bound thereto, at least one lanthanide metal chelate or at least one fluorophore, each acting as either an energy donor or an energy acceptor. Also disclosed are a method and kit for its use.
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Citations
53 Claims
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1. A nucleic acid hybridization composition comprising:
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an oligo- or polynucleotide having directly or indirectly bound thereto at least one lanthanide metal or at least one fluorophore, each acting as either an energy donor or an energy acceptor; a solid matrix having at least one surface to which is attached a first intercalator capable of capturing a double-stranded nucleic acid; a second intercalator, that may or may not comprise at least one fluorophore, said second intercalator or said fluorophore, each acting as either an energy donor or an energy acceptor wherein upon hybridization of said oligo- or polynucleotide to a complementary oligo- or polynucleotide, said energy donor and said energy acceptor are within proximate distance of each other such that energy from said energy donor is absorbed by said energy acceptor. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24)
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25. A nucleic acid hybridization composition comprising:
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an oligo- or polynucleotide having directly or indirectly bound thereto at least one lanthanide metal or at least one fluorophore, each acting as either an energy donor or an energy acceptor; a solid matrix having at least one surface to which is attached (i) a first intercalator capable of capturing a double-stranded nucleic acid and acting as either an energy donor or an energy acceptor at a characteristic fluorescence emision; and
(ii) a second intercalator that may or may not comprise at least one fluorophore, wherein said second intercalator is capable of capturing double-stranded nucleic acid, and wherein said second intercalator or said fluorophore acts as either an energy donor or an energy acceptor at a characteristic fluorescence emission which is different from the characteristic fluorescence emission of said first intercalator.
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26. A nucleic acid hybridization assay process for detecting in a sample the presence of a nucleic acid sequence of interest, which process comprises:
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(a) contacting a sample suspect of containing said nucleic acid sequence of interest with an oligo- or polynucleotide hybridizable therewith, said nucleic acid sequence of interest being present in single-stranded form or having been rendered at least partially single-stranded, and said oligo- or polynucleotide having directly or indirectly bound thereto a lanthanide metal or a fluorophore, each independently acting as either an energy donor or an energy acceptor; (b) permitting hybridization of said nucleic acid sequence of interest and said oligo- or polynucleotide to form a complex; (c) contacting said complex with; (i) a solid matrix having at least one surface to which is attached a first intercalator capable of capturing double-stranded nucleic acid, and (ii) a second intercalator with or without a fluorescent compound attached thereto, each independently acting as either an energy donor or an energy acceptor, the fluorescence emissions of said first and second intercalators being characterized by different wavelengths; wherein upon hybridization of said oligo- or polynucleotide to said nucleic acid sequence of interest, said energy donor and said energy acceptor are within proximate distance of each other such that energy from said energy donor is absorbed by said energy acceptor; and (d) detecting any energy emitted from said energy acceptor. - View Dependent Claims (27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52)
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42. The process of 26 wherein the double-stranded nucleic acid capturable by said first intercalator is selected from the group consisting of double-stranded DNA, double-stranded RNA and DNA-RNA hybrids.
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53. A nucleic acid hybridization assay kit comprising, in packaged combination, reagents for detecting in a sample the presence of a nucleic acid sequence of interest, including the following:
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a first container having therein a plurality of solid matrices having bound to at least one surface thereof a first intercalator capable of capturing double-stranded nucleic acid; a second container having therein an oligo- or polynucleotide hybridizable with said nucleic acid sequence and having directly or indirectly bound thereto at least one lanthanide metal or at least one fluorophore each acting as either an energy donor or an energy acceptor; and at least one additional container having therein a second intercalator with or without at least one fluorophore attached thereto, each acting as either an energy donor or an energy acceptor, wherein the fluorescence emissions of said first and second intercalators are characterized by different wavelengths.
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Specification