Compositions and methods for performing hybridizations with separate denaturation of the sample and probe
AssigneeDako Denmark A/S
InventorSteen Hauge Matthiesen
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Abstract
The invention provides methods and compositions for separately denaturing a probe and target in hybridization applications. The invention may, for example, eliminate the use of, or reduce the dependence on formamide in hybridization applications. Compositions for use in the invention include an aqueous composition comprising at least one polar aprotic solvent in an amount effective to denature double-stranded nucleotide sequences.
Claims61
A method of hybridizing nucleic acid sequences comprising:
- providing a first nucleic acid sequence within a sample having a preserved cell morphology, with a first composition comprising at least one polar aprotic solvent in an amount effective to denature a double-stranded nucleotide sequence while preserving cell morphology, and at least 10% dextran sulfate,
- providing a second nucleic acid composition comprising a second nucleic acid sequence and a second aqueous composition comprising at least one denaturing agent in an amount effective to denature double-stranded nucleotide sequences, and
- combining the first and the second nucleic acid sequence compositions for at least a time period sufficient to hybridize the first and second nucleic acid sequences such that the cell morphology is preserved,
- wherein the polar aprotic solvent is not dimethyl sulfoxide (DMSO).
A method of hybridizing nucleic acid sequences comprising:
- providing a first nucleic acid sequence, within a sample having a preserved cell morphology, with a first composition comprising at least one polar aprotic solvent in an amount effective to denature a double-stranded nucleotide sequence while preserving cell morphology, and at least 10% dextran sulfate, and
- combining said first nucleic acid composition with a second nucleic acid composition comprising a second nucleic acid sequence and at least one denaturing agent in an amount effective to denature double-stranded nucleotide sequences for at least a time period sufficient to hybridize the first and second nucleic acid sequences such that the cell morphology is preserved,
- wherein the polar aprotic solvent is not dimethyl sulfoxide (DMSO).
A method of hybridizing nucleic acid sequences comprising:
- providing a first nucleic acid sequence within a sample having a preserved cell morphology, with a composition comprising at least one polar aprotic solvent in an amount effective to denature a double-stranded nucleotide sequence while preserving cell morphology, and at least 10% dextran sulfate, and
- combining said nucleic acid composition with a second nucleic acid sequence for at least a time period sufficient to hybridize the first and second nucleic acid sequences such that the cell morphology is preserved,
- wherein the polar aprotic solvent is not dimethyl sulfoxide (DMSO).
An aqueous composition for performing separate denaturation of a target in a hybridization application within a sample having a preserved cell morphology, said composition comprising at least one polar aprotic solvent in an amount effective to denature a double-stranded nucleotide sequence while preserving cell morphology and at least 10% dextran, wherein the polar aprotic solvent is not dimethyl sulfoxide (DMSO).
Method of denaturing a target within a sample having a preserved cell morphology comprising combining the target within the sample with a composition comprising between 1 and 90% (v/v) of at least one polar aprotic solvent and at least 10% dextran sulfate, wherein the polar aprotic solvent is not dimethyl sulfoxide (DMSO), wherein the at least one polar aprotic solvent denatures the target while preserving cell morphology.
Dependencies are inferred from the claim text.