Microcalorimetric detection of analytes and binding events

• US 20030059807A1
• Filed: 06/07/2002
• Published: 03/27/2003
• Est. Priority Date: 06/07/2001
• Status: Abandoned Application

• Current Assignee: Sigma-Aldrich Company (Sigma-Aldrich Corporation)
• Original Assignee: Proligo, LLC
• Inventors: Roach, Warren, Roach, Jeffrey Shawn, Wolter, Andreas
• Application Number:
  US10/165,854
• Publication Number:
  US 20030059807A1
• Time in Patent Office: Days
• US Class Current: 435/6
• CPC Class Codes: B01L 3/5027 by integrated microfluidic structures, i.e. dimensions of channels and chambers are such that surface tension forces are important, e.g. lab-on-a-chip B01L3/5023 takes precedence; micromixers B01F33/30; microreactors for synthesis B01J19/0093; microcapillary devices in general B81B1/00
• CPC Class Codes: B01L 3/5085 for multiple samples, e.g. microtitration plates
• CPC Class Codes: C12Q 1/6825 Nucleic acid detection involving sensors
• CPC Class Codes: C12Q 1/6837 using probe arrays or probe chips C12Q1/6874 takes precedence
• CPC Class Codes: C12Q 2527/101 Temperature
• CPC Class Codes: C12Q 2565/607 being a sensor, e.g. electrode
• CPC Class Codes: G01N 33/54373 involving physiochemical end-point determination, e.g. wave-guides, FETS, gratings

Abstract

The present invention comprises methods for detecting specific binding interactions through measuring the heat of binding generated when members of specific binding pairs interact with each other. The invention also comprises methods to detect analytes in a solution through measurement of the heat of binding or reaction generated from the interaction of the analytes with binding or reaction partners. In addition, the invention comprises detection devices that consist of spatially addressable arrays of thermistors, which are useful in the multi-parallel thermal analysis of samples. The analytical methods and devices described are particularly useful in the analysis of nucleic acids.

Claims

1. A method for detecting binding between a first member of a specific binding pair and a second member of a specific binding pair, said method comprising:

a) providing a detection device, wherein said detection device comprises an array of spatially addressable thermistors;

b) providing the first member of the specific binding pair, wherein said first member of the specific binding pair is closely associated with a spatially addressable thermistor in the detection device;

c) contacting the detection device with a sample containing one or more of the second member of the specific binding pair; and

d) detecting the binding between said binding pair members via thermal analysis.

2. The method of claim 1 wherein said binding pair is selected from the group consisting of complementary nucleic acids, antibody/antigen, ligand/receptor, enzyme/substrate and aptamer/target.

3. The method of claim 2 wherein the specific binding pair comprises chimeric molecules.

4. The method of claim 2 wherein said complementary nucleic acids are selected from the group consisting of DNA/DNA, DNA/RNA, DNA/LNA, DNA/siRNA and DNA/PNA.

5. The method of claim 2 wherein said binding is hybridization between the complementary nucleic acids.

6. The method of claim 1 wherein the first member of the specific binding pair is attached to the detection device via a covalent bond or a non-covalent bond.

7. The method of claim 1 wherein said first member of the specific binding pair is localized over the detection device in the form of a solution that is in close proximity to the thermistor of the detection device.

8. The method of claim 1 wherein said thermistor is a negative temperature coefficient (NTC) thermistor.

9. The method of claim 1 wherein each member of said binding pair contains a reactive moiety.

10. The method of claim 9 wherein said reactive moiety is selected from groups selected to maximize the heat of reaction.

11. The method of claim 9 wherein and said binding between said binding pair members is a covalent reaction between the reactive moieties.

12. The method of claim 9 wherein said binding between said binding pair members is via noncovalent interactions followed by covalent reaction between said reactive moieties on each member of said binding pair.

13. The method of claim 1 wherein the detection device provides a real time, digital profile of the binding between said binding pair members as is occurs in the detection device.

14. A method for detecting an analyte in a solution said method comprising:

a) providing a detection device, wherein said detection device is comprised of an array of spatially addressable thermistors;

b) providing a binding or reaction partner to said analyte, wherein said binding or reaction partner to the analyte is closely associated with a spatially addressable thermistor in the detection device;

c) contacting the detection device with a sample containing one or more analytes; and

d) detecting the binding or reaction between said analyte and its binding or reaction partner by thermal analysis.

15. The method of claim 14 wherein the binding or reaction partner and the analyte are selected from the group consisting of complementary nucleic acids, antibody/antigen, ligand/receptor, enzyme/substrate and aptamer/target.

16. The method of claim 15 wherein the binding or reaction partner and the analyte are comprised of chimeric molecules.

17. The method of claim 15 wherein said complementary nucleic acids are selected from the group consisting of DNA/DNA, DNA/RNA, DNA/LNA, DNA/siRNA and DNA PNA.

18. The method of claim 15 wherein said binding is hybridization between the complementary nucleic acids.

19. The method of claim 14 wherein the binding or reaction partner is attached to the detection device via a covalent bond or a non-covalent bond.

20. The method of claim 14 wherein said the binding or reaction partner is localized over the detection device in the form of a solution that is in close proximity to the thermistors of the detection device.

21. The method of claim 14 wherein said thermistors are negative temperature coefficient (NTC) thermistors.

22. The method of claim 14 wherein the binding or reaction partner and the analyte each contain a reactive moiety.

23. The method of claim 22 wherein said reactive moiety is selected from groups selected to maximize the heat of reaction.

24. The method of claim 22 wherein and said reaction between the analyte and its binding or reaction partner is a covalent reaction between the reactive moieties.

25. The method of claim 22 wherein said reaction between the analyte and its binding or reaction partner is via noncovalent interactions followed by covalent reaction between said reactive moieties.

26. The method of claim 14 wherein two molecules jointly provide the binding or reaction partner of an analyte and wherein at least one of the two binding or reactions is closely associated with the thermistors of the detection device.

27. The method of claim 26 wherein at least one of the two molecules and the analyte each contain a reactive moiety.

28. The method of claim 27 wherein said reaction is a chemical ligation.

29. The method of claim 14 wherein the binding between the analyte and its binding partner is used to distinguish between perfectly complementary sequences and non-complementary sequences in which the non-complementary elements may comprise one or more elements of the mismatched sequence.

30. The method of claim 14 wherein the binding between the analyte and its binding partner comprises part of an enzymatic amplification reaction.

31. The method of claim 30 wherein said enzymatic amplification is a polymerase chain reaction or a primer extension reaction.

32. The method of claim 14 wherein the detection device provides a real time, digital profile of the binding or reaction between the analyte and its binding or reaction partner.

33. A detection device comprised of an array of addressable thermistors, wherein each of said addressable thermistors is closely associated to either a first member of a specific binding pair or to a binding or reaction partner to an analyte.

34. The detection device of claim 33 wherein said thermistors are negative temperature coefficient (NTC) thermistors.

35. The detection device of claim 33 wherein the array of addressable thermistors is further comprised of reservoirs that encompass the thermistors.

36. The detection device of claim 35 wherein said reservoir is a multi-well microtitre plate.

37. The detection device of claim 33 wherein the array of addressable thermistors is integrated onto a planar surface.

38. The detection device of claim 37 wherein said planar surface is a microelectronic chip.

39. The detection device of claim 38 wherein said microelectronic chip is a silicon wafer or equivalent material thereof.

40. The detection device of claim 33 wherein the means of association of the binding pair or reaction partners with the thermistor is selected from covalent or non-covalent attachment, or spatial localization in the form of a solution that is in close proximity to the thermistors of the detection device.

41. The detection device of claim 33 wherein said array of thermistors is housed in a chamber to control both temperature and humidity.

42. The detection device of claim 33 wherein each of said thermistors is interfaced to a processing unit.

43. The detection device of claim 33 further comprising means to provide a real time signal output.

44. A method for preparing a detection device comprising:

a) providing a solid support; and

b) associating at least two thermistors with the solid support in an addressable array, connecting the thermistors and further associating at least one spot of a first member of a specific binding pair or reaction partner of known composition surrounding and in close proximity to the thermistors, wherein each thermistor is connected to a signal processor.

45. The method of claim 44 wherein said thermistors are negative temperature coefficient (NTC) thermistors.

46. The method of claim 44 wherein said solid support is selected from the group consisting of Langmuir-Bodgett films, functionalized glass, germanium, silicon, silicon carbide, PTFE, polystyrene, gallium arsenide, gold and silver.

47. The method of claim 44 wherein the array of addressable thermistors is further comprised of reservoirs that encompass the thermistors.

48. The method of claim 47 wherein said reservoir is a multi-well microtitre plate.

49. The method of claim 44 wherein the array of addressable thermistors is integrated onto a planar surface or a spherical surface.

50. The method of claim 49 wherein said planar surface is a microelectronic chip.

51. The method of claim 50 wherein said microelectronic chip is a silicon wafer.

53. The method of claim 44 wherein said array of thermistors is housed in a chamber to control both temperature and humidity.

54. The method of claim 44 wherein each of said thermistors is interfaced to a processing unit.

55. The method of claim 44 further comprising providing means to produce a real time signal output.

56. The method of claim 44 wherein said thermistors are connected via a bridged circuit configuration

52. The method 44 wherein the means of association of the binding pair or reaction partners with the thermistor is selected from a covalent or non-covalent attachment, or sapatial localization in the form of a solution that is in close proximity to the thermistors of the detection device.

57. An instrument for detecting a thermal event comprising:

a) a detection device comprised of an array of addressable thermistors, wherein each of said addressable thermistors is closely associated to either a first member of a specific binding pair or to a binding or reaction partner to an analyte and wherein each of said thermistors is interfaced to a processing unit; and

b) a delivery device comprised of an injector, for introducing a substance to the detection device.

58. The instrument of claim 57 further comprising an environmental control chamber to house the delivery device and detection device.

59. The instrument of claim 57 wherein said delivery device is an array of capillaries, quills or microdispensing nozzles mounted on a robotic device capable of controlled motion in the x, y and z direction.