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US 6,270,965 B1Granted
Integrated method and system for amplifying and for detecting nucleic acids
AssigneeRoche Diagnostics GmbH
InventorsJörg Kleiber, Karim Tabiti, Gregor Sagner
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Abstract
The present invention concerns a method and a device which enables an integrated amplification and detection of nucleic acids.
Claims16
A method for detection of a nucleic acid comprising:
- (a) providing a sample in which it is intended to detect a nucleic acid,
- (b) amplifying the nucleic acid in a closed device in the presence of at least one nucleic acid primer which is specific for the nucleic acid to be detected and at least one capture probe which is specific for the nucleic acid to be detected, the capture probe being immobilized on at least one inner surface of the device and selected in such a way that a hybrid of nucleic acid and capture probe has a lower melting temperature than a hybrid of nucleic acid and primer and wherein the amplification is carried out under such conditions that a hybrid of nucleic acid and primer is stable whereas a hybrid of nucleic acid and capture probe is unstable, thereby forming an amplification product,
- (c) binding the amplification product to the immobilized capture probe, and
- (d) detecting the binding of the amplification product to the capture probe by evanescence-induced fluorescence,
- wherein said closed device is closed during the entire amplifying and detecting procedure.
2 more elements
A reagent kit for detection of nucleic acids comprising:
- (a) a closed device for the amplification and detection of nucleic acids containing at least one capture probe immobilized on at least one inner surface thereof,
- (b) at least one nucleic acid primer,
- (c) optionally deoxyribonucleotides,
- (d) optionally a suitable enzyme for extending the nucleic acid primer, wherein the capture probe is selected such that a hybrid of the nucleic acid to be detected and the capture probe has a lower melting temperature than a hybrid of the nucleic acid to be detected and primer,
- (e) a means for sealing said device during said amplification and detection, and
- (f) a means for detecting binding of the nucleic acids to the capture probe by evanescence-induced fluorescence.
3 more elements
A method for detection of a nucleic acid comprising:
- (a) providing a sample in which it is intended to detect a nucleic acid,
- (b) amplifying the nucleic acid in a closed device in the presence of at least one nucleic acid primer which is specific for the nucleic acid to be detected and at least one capture probe which is specific for the nucleic acid to be detected, wherein the capture probe is immobilized on at least one optically transparent inner surface of the closed device and fluorescent labels are incorporated into an amplification product, and
- (c) detecting the nucleic acid by binding the amplification product to the immobilized capture probe by means of evanescence-induced fluorescence,
- wherein said closed device is closed during the entire amplifying and detecting procedure.
1 more element
A reagent kit for detection of nucleic acids comprising:
- (a) a closed device for the amplification and detection of nucleic acids containing at least one capture probe immobilized on at least one inner surface thereof,
- (b) at least one nucleic acid primer,
- (c) optionally deoxyribonucleotides,
- (d) optionally an enzyme suitable for the extension of the nucleic acid primer,
- (e) a fluorescent label,
- (f) optionally a means for detection of amplification products by evanescence-induced fluorescence,
- (g) a means for sealing said device during said amplification and detection.
4 more elements
Dependencies are inferred from the claim text.