Methods and articles for strand-specific polynucleotide detection with cationic multichromophores
First Claim
1. An assay method comprising:
- providing a sample that is suspected of containing a target polynucleotide;
providing a polycationic multichromophore that comprises a conjugated polymer or dendrimer, wherein said multichromophore has a detectable excitation emission;
providing a sensor that is a polynucleotide that is complementary to the target polynucleotide, wherein said sensor is conjugated to a substrate and is not conjugated to a fluorescently detectable label that can receive energy from an excited state of the multichromophore;
contacting the substrate with the sample;
contacting the substrate with the multichromophore under conditions wherein the multichromophore can bind to the sensor and/or target polynucleotide, if present; and
detecting whether target has bound to the sensor.
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Abstract
The invention further relates to polycationic multichromophores, which may be conjugated polymers, and methods, articles and compositions employing them as described herein. In some aspects, the invention relates to methods, articles and compositions for the detection and analysis of biomolecules in a sample. Provided assays include those determining the presence of a target biomolecule in a sample or its relative amount, or the assays may be quantitative or semi-quantitative. The methods can be performed on a substrate. The methods can be performed in an array format on a substrate, which can be a sensor. In some embodiments, detection assays are provided employing sensor biomolecules that do not comprise a fluorophore that can exchange energy with the cationic multichromophore. In some aspects biological assays are provided in which energy is transferred between one or more of the multichromophore, a label on the target biomolecule, a label on the sensor biomolecule, and/or a fluorescent dye specific for a polynucleotide, in all permutations. The multichromophore may interact at least in part electrostatically with the sensor and/or the target, and an increase in energy transfer with the polymer may occur upon binding of the sensor and the target. Other variations of the inventions are described further herein.
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Citations
48 Claims
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1. An assay method comprising:
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providing a sample that is suspected of containing a target polynucleotide;
providing a polycationic multichromophore that comprises a conjugated polymer or dendrimer, wherein said multichromophore has a detectable excitation emission;
providing a sensor that is a polynucleotide that is complementary to the target polynucleotide, wherein said sensor is conjugated to a substrate and is not conjugated to a fluorescently detectable label that can receive energy from an excited state of the multichromophore;
contacting the substrate with the sample;
contacting the substrate with the multichromophore under conditions wherein the multichromophore can bind to the sensor and/or target polynucleotide, if present; and
detecting whether target has bound to the sensor. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22)
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23. A kit for assaying a sample for a target polynucleotide comprising:
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a polycationic multichromophore that is a conjugated polymer and that can electrostatically interact with a polyanionic target polynucleotide;
a substrate comprising a sensor polynucleotide that is complementary to the target polynucleotide, wherein said sensor polynucleotide is not conjugated to a fluorescently detectable label that can receive energy from an excited state of the multichromophore. - View Dependent Claims (24)
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25. A detection method comprising:
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providing a sample that is suspected of containing a target biomolecule;
providing a chromophore that comprises a polymer or dendrimer, wherein said chromophore has a detectable excitation emission;
providing a sensor biomolecule capable of recognition pairing with the target biomolecule, wherein the sensor is conjugated to a substrate and is not conjugated to a fluorescently detectable label that can receive energy from an excited state of the chromophore;
contacting the substrate with the sample under conditions in which the sensor can pair with the target, if present;
contacting the substrate with the multichromophore under conditions wherein the multichromophore can bind to the sensor and/or target polynucleotide, if present;
detecting whether target has bound to the sensor. - View Dependent Claims (26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 44, 45, 46, 47, 48)
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42. A kit for assaying a sample for a target comprising:
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a chromophore that is a polymer and that can electrostatically interact with the target;
a substrate comprising a sensor capable of recognition pairing with the target, wherein the sensor is not conjugated to a fluorescently detectable label that can receive energy from an excited state of the chromophore.
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43. A detection method comprising:
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providing a sample that is suspected of containing a polyanionic target comprising single-stranded DNA;
providing a polycationic multichromophore that comprises a conjugated polymer or dendrimer and that electrostatically interacts with the target and has a detectable excitation emission when excited with light comprising a wavelength of about 488 nm;
providing a sensor comprising PNA capable of recognition pairing with the target and not stably associating with the multichromophore in the absence of the target, wherein the sensor is conjugated to a substrate and is not conjugated to a fluorescently detectable label that can receive energy from an excited state of the multichromophore;
contacting the substrate with the sample under conditions in which the sensor can pair with the target, if present;
contacting the substrate with the multichromophore under conditions wherein the multichromophore can bind to the sensor and/or target polynucleotide, if present;
detecting whether target has bound to the sensor.
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Specification