US5283174A
Expired / Lapsed
US
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Homogenous protection assay
Assignee: Gen Probe Inc
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Filing: 1990-11-08
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Granted: 1994-02-01
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Expires: 2010-11-08
Abstract
Abstract Improved homogenous diagnostic assay methods and labels for detecting an analyte in a medium when the analyte is a member of a specific binding pair. The methods and labels provide procedures for reducing background and increasing sensitivity. The binding partner of the analyte is labeled with a substance, the stability of which detectably changes whenever said analyte is bound as a member of the specific binding pair. In a closely related system, the analyte is labeled with a substance susceptible to differential degradation depending on whether or not the analyte is bound as a member of its specific binding pair. After incubation and selective degradation or chemical or biochemical alteration, the amount of analyte bound is detected by measuring either the stability change or the extent of degradation of the label. In a particular system, chemiluminescent acridinium ester labeled probes are used in a homogenous hybridization assay format for sensitively detecting the presence of complement any target polynucleotide sequences.
Inventors
- 👤 Lyle J. Arnold, Jr.
- 👤 Norman C. Nelson
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-11-06 17:24
CPC Classifications
G; G01; G01N; G01N33/00; G01N33/48; G01N33/50; G01N33/58; G01N33/582; C; C12; C12Q; C12Q1/00; C12Q1/68; C12Q1/6813; C12Q1/6816; C12Q1/6876; C12Q1/6883; C12Q1/6886; C12Q1/6888; C12Q1/689; G01N33/53; G01N33/5306; G01N33/531; G01N33/532; G01N33/533; G01N33/536; G01N33/542; C12Q2600/00; C12Q2600/158
Claims Analysis
Total Claims: 13
Claim 1
Independent Claim
193 words
A homogeneous assay method for determining the presence or amount of a first nucleic acid in a medium, comprising the steps of: contacting a free second nucleic acid with said medium to form a hybridization mixture, wherein said free second nucleic acid is able to hybridize with said first nucleic acid under hybridizing conditions to form a binding pair, and wherein said second nucleic acid is chemically bonded to an N-acridinium ester which is selectively susceptible to chemical degradation by a chemical selected from the group consisting of an acid, a base, and an oxidizing agent, to form an altered N-acridinium ester dependent upon whether said second nucleic acid is free or forms part of said binding pair; treating said hybridization mixture with said chemical such that N-acridinium ester chemically bonded to free second nucleic acid is degraded to form altered N-acridinium ester and N-acridinium ester present in a binding pair is undegraded; and determining the amount of undegraded N-acridinium ester remaining after said treating step, without physically separating any said free second nucleic acid from any said binding pair, as a measure of the presence or amount of said first nucleic acid.
Claim 2
Dependent Claim
193 words
A homogeneous assay method for determining the presence or amount of a first nucleic acid in a medium, comprising the steps of: contacting a free second nucleic acid with said medium to form a hybridization mixture, wherein said free second nucleic acid is able to hybridize with said first nucleic acid under hybridizing conditions to form a binding pair, and wherein said second nucleic acid is chemically bonded to an N-acridinium ester which is selectively susceptible to chemical degradation by a chemical selected from the group consisting of an acid, a base, and an oxidizing agent, to form an altered N-acridinium ester dependent upon whether said second nucleic acid is free or forms part of said binding pair; treating said hybridization mixture with said chemical such that N-acridinium ester chemically bonded to free second nucleic acid is degraded to form altered N-acridinium ester and N-acridinium ester present in a binding pair is undegraded; and determining the amount of undegraded N-acridinium ester remaining after said treating step, without physically separating any said free second nucleic acid from any said binding pair, as a measure of the presence or amount of said first nucleic acid.
Claim 3
Dependent Claim
20 words
The method of claim 1 wherein determining the amount of label comprises the addition of H 2 O 2 .
Claim 4
Dependent Claim
11 words
The method of claim 1 wherein said chemical is an acid.
Claim 5
Dependent Claim
11 words
The method of claim 1 wherein said chemical is a base.
Claim 6
Dependent Claim
12 words
The method of claim 1 wherein said chemical is an oxidizing agent.
Claim 7
Dependent Claim
12 words
The method of claim 3 wherein said acid is HNO 3 .
Claim 8
Dependent Claim
10 words
The method of claim 4 wherein said base is NaOH.
Claim 9
Dependent Claim
19 words
The method of claim 1 wherein said method allows detection of 10 -16 Mol of said first nucleic acid.
Claim 10
Dependent Claim
162 words
The method of claim 1, in which said N-acridinium ester comprises the chemical formula ##STR11## wherein is selected from the group consisting of O, N, S, halogen, phosphorus, boron, and arsenic; Y is selected from the group consisting of O, S, or NH; R 1 is selected from the group consisting of alkyl, alkenyl, aryl, alkoxy, and aryloxy, or wherein R 1 is absent when X is a halogen; R 2 is selected from the group consisting of H, alkyl, alkenyl, aryl, alkoxy and aryloxy when X is N, or wherein R 2 is absent when X is not N; R 3 is selected from the group consisting of H, amino, hydroxy, thiol, halogen, nitro, amido, acetyl, alkyl, aryl, alkenyl, alkoxy, and aryloxy; R 4 is selected from the group consisting of alkyl, alkenyl, and aryl; and wherein at least one of R 1 , R 2 , R 3 , or R 4 contains a reactive site which allows chemical conjugation.
Claim 11
Dependent Claim
11 words
The method of claim 9 wherein said chemical is an acid.
Claim 12
Dependent Claim
11 words
The method of claim 9 wherein said chemical is a base.
Claim 13
Dependent Claim
12 words
The method of claim 9 wherein said chemical is an oxidizing agent.
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USPTO Annuity & Maintenance Windows
| Annuity Stage | Window Opens | Payment Deadline | Surcharge Period Until | Estimated Fee | Status |
|---|---|---|---|---|---|
| 3.5 Years (Year 4) | 1997-01-31 | 1997-08-02 | 1998-02-01 | $2000.00 | Window Closed / Lapsed |
| 7.5 Years (Year 8) | 2001-01-31 | 2001-08-02 | 2002-02-01 | $3760.00 | Window Closed / Lapsed |
| 11.5 Years (Year 12) | 2005-01-31 | 2005-08-02 | 2006-02-01 | $7700.00 | Window Closed / Lapsed |