Device and method for multiple analyte detection
View Patent ↗The invention is directed to a method and device for simultaneously testing a sample for the presence, presence, and/or amounts of one or more of a plurality of selected analytes. The invention includes, in one aspect, device for detecting or quantitating a plurality of different analytes in a liquid sample. Each chamber may include analyte-specific reagent effective to react with a selected analyte that may be present in the sample, and detection means for detecting the signal. Also disclosed are methods utilizing the device.
1. A method of detecting or quantitating a target nucleic acid sequence in a liquid sample, comprising:
pre-loading a first unlabeled oligonucleotide into a chamber;
pre-loading a second unlabeled oligonucleotide into the chamber;
introducing a liquid sample comprising a target nucleic acid sequence into the chamber, after the pre-loading of the first unlabeled oligonucleotide and of the second unlabeled oligonucleotide;
mixing the liquid sample with the first unlabeled oligonucleotide and the second unlabeled oligonucleotide, in the chamber, such that the first unlabeled oligonucleotide and the second unlabeled oligonucleotide are not immobilized in the chamber;
reacting the target nucleic acid sequence with both the first unlabeled oligonucleotide and the second unlabeled oligonucleotide in the chamber, to form a reaction product in the chamber;
reacting the reaction product, in the chamber, with a non-immobilized optically detectable detection reagent that is specific for the target nucleic acid sequence, wherein the non-immobilized optically detectable detection reagent is pre-loaded into the chamber before the introducing of the sample; and
detecting the non-immobilized optically detectable detection reagent to detect or quantitate the target nucleic acid sequence.
2. The method of claim 1 , wherein reacting the target nucleic acid sequence with both the first unlabeled oligonucleotide and the second unlabeled oligonucleotide in the chamber, to form a reaction product in the chamber, comprises at least one of a polymerase chain reaction, a ligase chain reaction, an oligonucleotide ligation assay, and a hybridization assay.
3. The method of claim 1 , wherein the first unlabeled oligonucleotide and the second unlabeled oligonucleotide comprise a primer pair suitable for amplifying the target nucleic acid sequence.
4. The method of claim 1 , wherein at least one of the first unlabeled oligonucleotide and the second unlabeled oligonucleotide comprises a sequence that is complementary to at least a portion of the target nucleic acid sequence.
5. The method of claim 1 , wherein the reaction product is detected through an optically transparent window in the chamber.
6. The method of claim 1 , wherein the target nucleic acid sequence is introduced into the chamber by vacuum action.
7. The method of claim 1 , wherein at least one of the pre-loaded first unlabelled oligonucleotide and the pre-loaded second unlabelled oligonucleotide are dried in the chamber prior to the mixing.
8. The method of claim 1 , wherein the optically detectable detection reagent comprises a fluorescent dye.
9. The method of claim 1 , wherein the optically detectable detection reagent comprises a fluorescer dye and a quencher dye.
10. The method of claim 1 , wherein at least one of the first unlabeled oligonucleotide and the second unlabeled oligonucleotide comprises a polynucleotide analog.
11. The method of claim 1 , wherein the first unlabeled oligonucleotide and the second unlabeled oligonucleotide bind to adjacent, contiguous regions of the target nucleic acid sequence.