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US 9,388,456 B2Granted
Compositions and methods for performing a stringent wash step in hybridization applications
AssigneeDako Denmark A/S
InventorSteen Hauge Matthiesen
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Abstract
The invention provides methods and compositions for performing a stringent wash step in a hybridization assay between at least one molecule and a target. The invention may, for example, eliminate the use of, or reduce the dependence on formamide in the stringent wash step. Compositions for use in the invention include an aqueous composition comprising at least one polar aprotic solvent in an amount effective to denature non-complementary sequences in a hybridization product.
Claims15
A method for performing a stringent wash step in a hybridization application comprising:
- a) providing a hybridization product comprising a first nucleic acid sequence hybridized to a second nucleic acid sequence within a cell having a sample morphology,
- b) providing an aqueous composition for performing a stringent wash, said composition comprising at least one polar aprotic solvent in an amount effective to denature non-complementary sequences in a hybridization product, a hybridization solution, and 5%-40% dextran sulfate, wherein the polar aprotic solvent is not dimethyl sulfoxide (DMSO) and has lactone, sulfone, sulfite, or carbonate functionality, and
- c) combining the hybridization product and the aqueous composition at a temperature of 82° C. or less for at least a time period sufficient to denature any non-complementary binding between the first and second nucleic acid sequences within the cell such that the sample morphology is preserved.
Dependencies are inferred from the claim text.