US 5,514,550 AGranted

Nucleic acid test article and its use to detect a predetermined nucleic acid

AssigneeJohnson & Johnson Clinical Diagnostics, Inc.

InventorsJohn B. Findlay, Janice M. Mayer, Marlene M. King, Fred T. Oakes, Chu-an Chang, Corey H. Levenson

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Abstract

A nucleic acid test article can be used to detect a targeted nucleic acid found in a specimen. The test article includes a substrate having two opposing surfaces and a water-insoluble nucleic acid probe attached in a distinct zone of one of the surfaces. The probe is prepared from a water-insoluble particle to which is covalently attached an oligonucleotide which is complementary to the targeted nucleic acid. Substantially none of the probe is embedded within the surface of the substrate. Particularly useful test articles have a multiplicity of water-insoluble probes located in distinct zones on one of the substrate surfaces. These probes are useful for the detection of a multiplicity of targeted nucleic acids, particularly after amplification by polymerase chain reaction.

Claims14

Dependent claims

A method for the detection of one or more predetermined nucleic acids, said method comprising:

A. amplifying one or more predetermined nucleic acids found in a specimen in the presence of complementary primers, deoxyribonucleotide triphosphates and a polymerization agent, at least one of said primers being detectably labeled, to provide one or more detectably labeled, amplified predetermined nucleic acids,
B. contacting said one or more detectably labeled, amplified predetermined nucleic acids with a nucleic acid test article comprising a substrate selected from the group consisting of a nonporous coated paper, a nonporous polymeric film, and a nonporous, noncoated paper, and having affixed in each of at least two distinct areas on said substrate, a water-insoluble nucleic acid probe, said nucleic acid test article being disposed in a self-contained test device,
each of said probes comprising an oligonucleotide complementary to a nucleic acid sequence of said one or more detectably labeled, amplified predetermined nucleic acids, which oligonucleotides are covalently attached to polymeric particles which are affixed predominantly on the outer surface of said substrate,
to form an immobilized hybridized product of each of said one or more detectably labeled, amplified predetermined nucleic acids and said water-insoluble probe in each of said distinct areas of said substrate,
C. washing said substrate to separate nonimmobilized materials from each immobilized product, and
D. detecting each immobilized product in each of said distinct areas of said substrate as a detection of the amount of one or more predetermined nucleic acids in said specimen,
wherein step C is carried out within about 4 minutes.

4 more elements

A method for the detection of one or more nucleic acid sequences of HIV-I DNA, said method comprising:

A. amplifying one or more nucleic acid sequences of HIV-I DNA found in a biological specimen in the presence of complementary primers, deoxyribonucleotide triphosphates and a polymerase isolated from a Thermus bacterial species, at least one of said primers being detectably labeled, to provide one or more detectably labeled, amplified nucleic acid sequences of HIV-I DNA,
B. contacting said one or more detectably labeled, amplified nucleic acid sequences of HIV-I DNA with a nucleic acid test article comprising a substrate selected from the group consisting of a nonporous coated paper, a nonporous polymeric film, and a nonporous, noncoated paper, having affixed in each of at least two distinct areas on said substrate, a water-insoluble nucleic acid probe, said nucleic acid test article being disposed in a self-contained test device,
each of said probes comprising an oligonucleotide complementary to said one or more detectably labeled, amplified nucleic acid sequences of HIV-I DNA, which oligonucleotides are covalently attached to polymeric particles having an average diameter of from about 0.1 to about 5 .mu.meters which are affixed predominantly on the outer surface of said substrate,
to form an immobilized hybridized product of each of one or more of said detectably labeled, amplified nucleic acid sequences of HIV-I DNA and said water-insoluble probe in each of said distinct areas of said substrate,
C. washing said substrate to separate nonimmobilized materials from each immobilized product, and
D. detecting each immobilized product in each of said distinct areas of said substrate as a detection of one or more nucleic acid sequences of HIV-I DNA in said specimen,
wherein step C is carried out within about 4 minutes.

4 more elements

Dependencies are inferred from the claim text.

Facts

Application: 08/300,297Filed 2 Sep 1994 · continuation
Priority: 3 Feb 1989
Granted: 7 May 1996
Term ends: Not estimatedThe source records don’t support an estimate.

Classification4 codes

C12Q 1/703
For retroviruses: viruses associated with AIDS
C12Q 1/6834
Hybridisation assays: enzymatic or biochemical coupling of nucleic acids to a solid phase
C12Q 1/6876
Involving nucleic acids: nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
C12Q
Measuring or testing processes involving enzymes, nucleic acids or microorganisms