Claims
- 1. A device for carrying out a plurality of different reactions in a single bulk-phase reaction medium, comprising:
means defining an elongate or planar channel and a port for introducing such bulk-phase medium into the channel, a plurality of discrete reaction regions within the channel, and a reaction-specific reagent releasably carried on a wall portion of each reaction region, for reacting in solution with one or more reagents in the bulk-phase medium, when such medium is introduced into the channel, to effect a selected solution-phase reaction in each region, where the channel is dimensioned to substantially prevent convective fluid flow among the reaction regions during such reactions.
- 2. The device of claim 1, wherein said channel defining means defines a one-dimensional channel having a substantially uniform cross-section along its length, channel width and depth dimensions between about 20-800 microns, and the reaction regions are submicroliter in volume.
- 3. The device of claim 1, wherein said channel defining means defines a channel having a plurality of radial bulges corresponding to the reaction regions, and connected in series by channel sections having channel width and depth dimensions between about 20-800 microns.
- 4. The device of claim 1, wherein said channel-defining means includes a pair of planar expanses that are separated from one another by a dimension between about 20-800 microns, and the reaction regions are submicroliter in volume.
- 5. The device of claim 1, for carrying out sequence-specific nucleic acid reactions involving target nucleic acid present in the bulk-phase medium, wherein the reaction-specific reagents are nucleic acid oligomer reagents releasably bound to the wall portions through duplex formation with immobilized complementary-sequence oligonucleotides, or via ligand attachment to an immobilized antiligand.
- 6. The device of claim 6, wherein each reaction region includes a capture nucleic acid immobilized on the associated wall portion and having a region-specific nucleic acid sequence, and wherein different-sequence nucleic acid oligomer reagents are hybridized with such capture nucleic acids.
- 7. The device of claim 6, for carrying out sequence-specific nucleic acid reactions selected from the group consisting of:
(a) polymerase extension reactions, wherein the reaction-specific reagents in each region include extension primers; (b) PCR reactions in the reaction regions, wherein the reaction-specific reagents in each region include one or more sets of PCR primers. (c) sequence-specific 5′ exonuclease reactions that result in the formation of a detectable product, wherein the reaction-specific reagent in each region include as an exonuclease substrate, an oligonucleotide having a selected nucleic acid sequence terminating in a detectably labeled 5′ nucleotide.
- 8. The device of claim 8, for use in carrying out 5′ exonuclease reactions, wherein detectably labeled 5′ nucleotides associated with different reaction regions are electrophoretically separable.
- 9. A device for carrying out simultaneous sequence-specific nucleic acid reactions on a plurality of DNA target segments (i) contained in a bulk-phase medium and (ii) having different nucleic acid sequences, comprising:
a substrate defining an elongate channel terminating at first and second ends, a lid covering the open channel to form an elongate closed channel terminating at first and second ports, a plurality of discrete reaction regions spaced along the length of said channel, between said ports, and in each reaction region, one or more region-specific nucleic acids releasably carried on a portion of that reaction region, where the region-specific nucleic acids are effective to bind to complementary sequence nucleic acid target segments contained in the bulk-phase medium, after such medium is introduced into the channel, and the channel design substantially prevents convective fluid flow among the reaction regions in the channel, whereby the region-specific nucleic acids are largely confined to the associated region during such reaction.
- 10. The device of claim 10, wherein each reaction region includes a capture nucleic acid immobilized on the associated wall portion and having a region-specific nucleic acid sequence, and wherein different-sequence nucleic acid oligomer reagents are hybridized with such capture nucleic acids.
- 11. The device of claim 10, for or carrying out sequence-specific nucleic acid reactions selected from the group consisting of:
(a) polymerase extension reactions, wherein the reaction-specific reagents in each region include extension primers; (b) PCR reactions in the reaction regions, wherein the reaction-specific reagents in each region include one or more sets of PCR primers, (c) sequence-specific 5′ exonuclease reactions that result in the formation of a detectable product, wherein the reaction-specific reagent in each region include as an exonuclease substrate, an oligonucleotide having a selected nucleic acid sequence terminating in a detectably labeled 5′ nucleotide.
- 12. The device of claim 10, wherein said substrate is designed to be placed in a centrifugation apparatus, such that centrifugation of the device is effective to cause liquid medium introduced at one port to fill the channel, or liquid medium contained within the channel to be expelled therefrom.
- 13. A method for simultaneously carrying out a plurality of different reactions that involve both common and reaction-specific reagents, comprising:
filling a channel having (i) means defining an elongate or planar channel and a port for introducing a liquid medium into the channel, and (ii) a reaction-specific reagent releasably carried on a wall portion of each reaction region, for reacting in solution with one or more reagents in the bulk-phase medium, when such medium is introduced into the channel, to effect a selected solution-phase reaction in each region, and by said filling, and with release of reaction-specific reagent from the wall portion in each reaction region, simultaneously promoting reactions involving reagents provided in the bulk phase and the reaction-specific reagents in each of the reaction regions.
- 14. The method of claim 14, wherein after the completion of said reactions, the medium is removed from the device for analysis or processing of the plurality of reaction products.
- 15. The method of claim 14, for carrying out simultaneous PCR reactions on a plurality of different DNA targets contained in the bulk-phase medium, wherein said reaction-specific reagents in the different reaction regions include PCR primers designed to hybridize with and amplify different, selected regions of the DNA targets.
- 16. The method of claim 15, wherein promoting said reaction includes successively heating and cooling the device, under conditions effective to produce PCR amplicons.
- 17. A method for carrying out a plurality of simultaneous sequence-specific nucleic acid reactions on a plurality of DNA target segments (i) contained in a bulk-phase medium and (ii) having different nucleic acid sequences, comprising
adding to device having (i) means defining an elongate channel and a port for introducing a liquid medium into the channel, and (ii) region-specific capture nucleic acids immobilized on channel wall portions at a plurality of discrete reaction regions contained within and along the length of the channel, a solution containing a plurality of different-sequence nucleic acid reagents, each having a capture portion effective to hybridize to one of the capture nucleic acids and a reaction portion effective to hybridize to one of the target DNA sequences in the bulk phase medium, under DNA hybridization conditions, thereby to localize selected nucleic acid reagents at selected reactions regions in the channel, filling said channel with such bulk phase medium, and simultaneously promoting reactions involving target segments contained in the bulk phase medium and such region-specific nucleic acid reagents, by causing release of the nucleic acid reagents from the associated reaction-region wall portions.
- 18. The method of claim 17, which further includes capturing reaction in each reaction region, by hybridization of reaction product to the immobilized capture nucleic acids.
- 19. A method for performing a plurality of affinity determinations to determine the biological activity of candidate compounds employing an elongated channel having a cross-section in the range of about 10 um2 to about 4 mm2 and a plurality of sites at which are non-diffusively bound a first component of said affinity determination, wherein each site is bordered by a source trench and a drain trench for moving components of said affinity determination to and away from said site, said affinity determination comprising the binding of a candidate compound to an enzyme and employing an enzyme substrate which results in a detectable product, said method comprising:
electrokinetically moving each of said candidate compounds from each of said source trenches to each of their respective sites and incubating the resulting mixture at each site, adding said substrate to said main channel, incubating the resulting mixture at each site, resulting in a detectable product, electrophoretically moving said detectable product from said site to said drain trench, and detecting said detectable product separate from other components of said affinity determination as a measure of said affinity determination, wherein the length of said site and the cross-section of said channel are chosen to have a reaction volume for said affinity determination of less than about 100 nL.
Parent Case Info
[0001] This application claims the benefit of U.S. Provisonal Application No. 60/183,626 filed Feb. 18, 2000, which is incorporated herein by reference in its entirety.
Provisional Applications (1)
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Number |
Date |
Country |
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60183626 |
Feb 2000 |
US |