Electrochemical assay for nucleic acids and nucleic acid probes
First Claim
1. An assay for determining a target nucleic acid sequence which comprises the steps of:
- (a) providing a probe material comprising,(i) a sequence of nucleic acids complementary to a given target sequence and,(ii) a first ligand chemically linked thereto and capable of a specific binding reaction with an antiligand;
(b) contacting the said probe material with an assay system comprising,(i) a mediator, enzyme, substrate system capable of transferring charge to an electrode surface when the enzyme is catalytically active, and(ii) a second ligand chemically linked to one of said mediator, enzyme or substrate, wherein the second ligand is capable of competing with the first ligand for binding with the antiligand, and (iii) said antiligand;
whereby said first ligand competes with said second ligand in a specific binding reaction with the antiligand, the extent of binding of said second ligand to said antiligand being reflected in the charge which said system transfers to said electrode surface;
(c) contacting the above system with a solution suspected of containing the said target sequence whereby the binding of any of the said target sequence present in the solution to the probe affects the availability of the first ligand to compete with said second ligand to bind said antiligand and therefore alters the rate of charge transfer to the electrode and(d) measuring said charge transfer and relating said measurement to the determination of said target nucleic acid sequence.
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Accused Products
Abstract
The present invention discloses an assay for nucleic acid which comprises the steps of;
(a) providing a probe material comprising;
(i) a sequence of nucleic acids complementary to a given target sequence and,
(ii) a first ligand chemically linked thereto and capable of a specific binding reaction with an antiligand;
(b) contacting the said probe material with an assay system comprising:
(i) a suitable mediator, enzyme, substrate system capable of transferring charge to an electrode surface when the enzyme is catalytically active, and;
(ii) a second ligand chemically linked to one of said mediator, enzyme or substrate, wherein the second ligand is capable of a competitive binding reaction with the antiligand, and;
(iii) the said antiligand, Whereby the said first ligand competes with the said second ligand in a specific binding reaction with the antiligand, and;
(c) contacting the above system with a solution suspected of containing the said target sequence whereby the binding of any of the said target sequence present to the probe affects the availability of the first ligand and therefore alters the rate of charge transfer to the electrode.
655 Citations
47 Claims
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1. An assay for determining a target nucleic acid sequence which comprises the steps of:
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(a) providing a probe material comprising, (i) a sequence of nucleic acids complementary to a given target sequence and, (ii) a first ligand chemically linked thereto and capable of a specific binding reaction with an antiligand; (b) contacting the said probe material with an assay system comprising, (i) a mediator, enzyme, substrate system capable of transferring charge to an electrode surface when the enzyme is catalytically active, and (ii) a second ligand chemically linked to one of said mediator, enzyme or substrate, wherein the second ligand is capable of competing with the first ligand for binding with the antiligand, and (iii) said antiligand; whereby said first ligand competes with said second ligand in a specific binding reaction with the antiligand, the extent of binding of said second ligand to said antiligand being reflected in the charge which said system transfers to said electrode surface; (c) contacting the above system with a solution suspected of containing the said target sequence whereby the binding of any of the said target sequence present in the solution to the probe affects the availability of the first ligand to compete with said second ligand to bind said antiligand and therefore alters the rate of charge transfer to the electrode and (d) measuring said charge transfer and relating said measurement to the determination of said target nucleic acid sequence. - 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, 25, 26, 27, 28)
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29. A method of assay for determining a target nucleic acid sequence comprising:
- (a) providing, (i) an electrode in contact with an enzyme and a substrate of the enzyme, and a mediator compound associated with the enzyme to transfer charge from the enzyme to the electrode as the conversion of substrate into product proceeds, and (ii) a probe nucleic acid sequence complementary to said target sequence which is linked to either the enzyme or the mediator compound;
(b) measuring a modification in the activity of the enzyme and the mediator compound and hence the quantity of charge being transferred to the electrode resulting from the binding of said target sequence to the probe nucleic acid sequence, and (c) relating said measurement to the determination of said target nucleic acid sequence. - View Dependent Claims (30, 31, 32, 33, 34, 35, 36, 37)
- (a) providing, (i) an electrode in contact with an enzyme and a substrate of the enzyme, and a mediator compound associated with the enzyme to transfer charge from the enzyme to the electrode as the conversion of substrate into product proceeds, and (ii) a probe nucleic acid sequence complementary to said target sequence which is linked to either the enzyme or the mediator compound;
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38. A nucleic acid probe for establishing the presence or copy number of a given target sequence in a single-stranded nucleic acid molecule, said nucleic acid probe being linked to
(a) either a metallocene mediator compound or an enzyme electrochemically coupled to an electrode by a metallocene mediator compound, and, (b) biotin, wherein binding of the probe nucleic acid to the target sequence modifies the electrochemical coupling between the metallocene mediator compound and the enzyme.
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41. A nucleic acid probe comprising:
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(a) a nucleic acid strand complementary to a target nucleic acid sequence, and (b) a metallocene mediator chemically linked thereto, wherein said mediator transfers charge from a catalytically active enzyme to an electrode. - View Dependent Claims (42, 43, 44, 45, 46, 47)
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Specification