EP0144914A2
Expired / Lapsed
EP
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Hybridization assay employing labeled pairs of hybrid binding reagents
Assignee: Bayer Corp
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Filing: 1984-11-29
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Granted: —
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Expires: 2004-11-29
Abstract
Abstract @ A nucleic acid hybridization assay wherein the hybrid formed with the probe has binding sites for two specific binding reagents, one of which comprises a first label and the other a second label, interaction of the first and second labels providing a detectable response which is measurably different when the two labeled reagents are both bound to the same hybrid compared to when the two labeled reagents are not so bound. The formation of the hybrid assay product brings the two labels within a proximate interaction distance of one another, e.g., as in the cases of sequential catalyst (enzyme) interaction and energy transfer. Since the labels provide a response which is distinguishable when they are associated with a hybridized probe, no separation step is required.
Inventors
- 👤 James P. Albarella
- 👤 Leslie H. Anderson Deriemer
- 👤 Robert J. Carrico
Bibliographic Details
Application Number
—
Jurisdiction
EP
Data Source
google_patents
Last Synchronized
2026-10-26 13:10
CPC Classifications
G; G01; G01N; G01N33/00; G01N33/48; G01N33/50; G01N33/53; G01N33/536; G01N33/542; C; C07; C07K; C07K16/00; C07K16/18; C12; C12Q; C12Q1/00; C12Q1/68; C12Q1/6804; C12Q1/6813; C12Q1/6816; C12Q1/6818
Claims Analysis
Total Claims: 17
Claim 1
Independent Claim
155 words
A method for detecting a particular polynucleotide sequence in a test sample comprising single stranded nucleic acids, comprising the steps of contacting the test sample with a nucleic acid probe comprising at least one single stranded base sequence that is substantially complementary to the sequence to be detected, and determining the resulting hybrids, characterized in that the hybrids are formed to have binding sites for two specific binding reagents, any hybrid that is formed is contacted with the two specific binding reagents, one of which comprises a first label and the other comprising a second label, interaction between the first and second labels providing a detectable response which is measurably different when the two labeled reagents are both bound to the same hybrid, compared to when the two labeled reagents are not so bound, and the detectable response is measured as a function of the presence of the sequence to be detected in the sample.
Claim 2
Independent Claim
155 words
A method for detecting a particular polynucleotide sequence in a test sample comprising single stranded nucleic acids, comprising the steps of contacting the test sample with a nucleic acid probe comprising at least one single stranded base sequence that is substantially complementary to the sequence to be detected, and determining the resulting hybrids, characterized in that the hybrids are formed to have binding sites for two specific binding reagents, any hybrid that is formed is contacted with the two specific binding reagents, one of which comprises a first label and the other comprising a second label, interaction between the first and second labels providing a detectable response which is measurably different when the two labeled reagents are both bound to the same hybrid, compared to when the two labeled reagents are not so bound, and the detectable response is measured as a function of the presence of the sequence to be detected in the sample.
Claim 3
Dependent Claim
58 words
The method of Claim 1 wherein the first label participates in a first chemical reaction that produces a diffusible product which is a participant in a second chemical reaction with the second label to produce a product that provides the detectable response, the first and second labels preferably being catalysts for the first and second chemical reactions, respectively.
Claim 4
Dependent Claim
17 words
The method of Claim 1 wherein the first and second labels participate in an energy transfer interaction.
Claim 5
Dependent Claim
16 words
The method of Claim 1 wherein one of the specific binding reagents is an antibody reagent.
Claim 6
Dependent Claim
58 words
The method of Claim 4 wherein the antibody reagent is one that is selective for binding DNA'RNA hybrids wherein one of the probe and the sequence to be detected is DNA and the other is RNA, or is one that is selective for binding RNA'RNA hybrids wherein both the probe and the sequence to be detected are RNA.
Claim 7
Dependent Claim
71 words
The method of Claim 4 wherein the antibody reagent is one that is selective for binding intercalation complexes wherein the duplexes formed in the assay comprise a nucleic acid intercalator bound thereto in the form of intercalation complexes, preferably the intercalator being chemically linked to the probe in its single stranded complementary region whereby upon hybridization with the sequence to be detected said intercalation complexes are formed in the resulting hybrid.
Claim 8
Dependent Claim
73 words
The method of either of Claim 5 or 6 wherein (i) the other specific binding reagent comprises the same antibody reagent, (ii) the nucleic acid probe comprises a specific binding ligand moiety and the other specific binding reagent is a binding partner for such ligand moiety, or (iii) the nucleic acid probe comprises a double stranded portion having a binding site for a protein and the other specific binding reagent is such protein.
Claim 9
Dependent Claim
27 words
The method of Claim 1 wherein the test sample comprises a biological sample which has been subject to conditions to release and denature nucleic acids present therein.
Claim 10
Dependent Claim
110 words
A reagent system for detecting a particular polynucleotide sequence in a test sample, comprising a nucleic acid probe comprising at least one single stranded base sequence that is substantially complementary to the sequences to be detected, characterized by comprising additionally first and second specific binding reagents capable of binding to hybrids formed between the sequence to be detected in the sample and the probe, such binding reagents comprising first and second labels, respectively, which interact with each other to provide a detectable response which is measurably different when the two labeled reagents are both bound to the same hybrid compared to when the two labeled reagents are not so bound.
Claim 11
Dependent Claim
110 words
A reagent system for detecting a particular polynucleotide sequence in a test sample, comprising a nucleic acid probe comprising at least one single stranded base sequence that is substantially complementary to the sequences to be detected, characterized by comprising additionally first and second specific binding reagents capable of binding to hybrids formed between the sequence to be detected in the sample and the probe, such binding reagents comprising first and second labels, respectively, which interact with each other to provide a detectable response which is measurably different when the two labeled reagents are both bound to the same hybrid compared to when the two labeled reagents are not so bound.
Claim 12
Dependent Claim
59 words
The reagent system of Claim 9 wherein the first label participates in a first chemical reaction that produces a diffusible product which is a participant in a second chemical reaction with the second label to produce a product that provides the detectable response, the first and second labels preferably being catalysts for the first and second chemical reactions, respectively.
Claim 13
Dependent Claim
18 words
The reagent system of Claim 9 wherein the first and second labels participate in an energy transfer interaction.
Claim 14
Dependent Claim
17 words
The reagent system of Claim 9 wherein one of the specific binding reagents is an antibody reagent.
Claim 15
Dependent Claim
59 words
The reagent system of Claim 12 wherein the antibody reagent is one that is selective for binding DNA'RNA hybrids wherein one of the probe and the sequence to be detected is DNA and the other is RNA, or is one that is selective for binding RNA'RNA hybrids wherein both the probe and the sequence to be detected are RNA.
Claim 16
Dependent Claim
72 words
The reagent system of Claim 12 wherein the antibody reagent is one that is selective for binding intercalation complexes wherein the duplexes formed in the assay comprise a nucleic acid intercalator bound thereto in the form of intercalation complexes, preferably the intercalator being chemically linked to the probe in its single stranded complementary region whereby upon hybridization with the sequence to be detected said intercalation complexes are formed in the resulting hybrid.
Claim 17
Dependent Claim
74 words
The reagent system of either of Claims 13 or 14 wherein (i) the other specific binding reagent comprises the same antibody reagent, (ii) the nucleic acid probe comprises a specific binding ligand moiety and the other specific binding reagent is a binding partner for such ligand moiety, or (iii) the nucleic acid probe comprises a double stranded portion having a binding site for a protein and the other specific binding reagent is such protein.
References Cited (Backward Citations)
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