Detection of nucleic acids from whole blood

• US 20070161015A1
• Filed: 10/04/2006
• Published: 07/12/2007
• Est. Priority Date: 10/05/2005
• Status: Active Grant

• Current Assignee: Affymetrix, Inc. (Thermo Fisher Scientific Incorporated)
• Original Assignee: Panomics, Inc.
• Inventors: Zheng, Zhi, Luo, Yuling, McMaster, Gary
• Granted Patent: US 7,927,798 B2
• Time in Patent Office: Days
• US Class Current: 435/6
• CPC Class Codes: C12Q 1/6816 characterised by the detection means C12Q1/6804 takes precedence
• CPC Class Codes: C12Q 1/6837 using probe arrays or probe chips C12Q1/6874 takes precedence
• CPC Class Codes: C12Q 2525/313 Branched oligonucleotides
• CPC Class Codes: C12Q 2537/125 Sandwich assay format
• CPC Class Codes: C12Q 2537/143 Multiplexing, i.e. use of multiple primers or probes in a single reaction, usually for simultaneously analyse of multiple analysis

Abstract

Methods of detecting one or more nucleic acids from whole blood or plasma are provided. The nucleic acids are captured on a solid support and detected. Compositions, kits, and systems related to the methods are also described.

Claims

1. A method of detecting at least a first target nucleic acid, the method comprising:

providing a sample comprising whole blood, which whole blood comprises peripheral blood cells;

lysing the peripheral blood cells to produce a lysate, which lysate comprises the first target nucleic acid;

contacting the first target nucleic acid with a first set of n capture extenders, wherein n is at least two, which first set of capture extenders is capable of hybridizing to the first target nucleic acid;

hybridizing the first target nucleic acid to the first set of capture extenders;

associating the first set of capture extenders with a solid support,

whereby hybridizing the first target nucleic acid to the first set of capture extenders and associating the first set of capture extenders with the solid support captures the first target nucleic acid on the solid support; and

detecting the presence of the first target nucleic acid on the solid support.

2. The method of claim 1, wherein the first target nucleic acid is an RNA.

3. The method of claim 1, wherein the first target nucleic acid is a DNA.

4. The method of claim 1, wherein contacting the first target nucleic acid with the first set of capture extenders comprises contacting the lysate with the first set of capture extenders.

5. The method of claim 1, wherein lysing the peripheral blood cells to produce a lysate, which lysate comprises the first target nucleic acid, comprises lysing the peripheral blood cells in the whole blood to produce a whole blood lysate, which whole blood lysate comprises the first target nucleic acid.

6. The method of claim 5, wherein contacting the first target nucleic acid with the first set of capture extenders comprises contacting the whole blood lysate with the first set of capture extenders.

7. The method of claim 1, wherein providing a sample comprising whole blood, which whole blood comprises peripheral blood cells, and lysing the peripheral blood cells to produce a lysate comprises:

applying the whole blood to a matrix to produce a blood spot;

drying the blood spot to produce a dried blood spot; and

contacting the dried blood spot with an aqueous solution to produce the lysate.

8. The method of claim 1, wherein the peripheral blood cells comprise white blood cells, one or more of which white blood cells comprises the first target nucleic acid.

9. The method of claim 1, the method comprising contacting the peripheral blood cells and/or the lysate with an exogenously supplied protease.

10. The method of claim 1, wherein n is at least three.

11. The method of claim 1, wherein n is at most 10.

12. The method of claim 1, wherein a first capture probe is bound to the solid support, and wherein associating the first set of capture extenders with the solid support comprises hybridizing the capture extenders to the first capture probe.

13. The method of claim 1, wherein the solid support is a substantially planar solid support.

14. The method of claim 1, wherein the solid support comprises a plurality of particles.

15. The method of claim 1, wherein the lysate comprises a second target nucleic acid, the method comprising:

contacting the second target nucleic acid with a second set of m capture extenders, wherein m is at least two, which second set of capture extenders is capable of hybridizing to the second target nucleic acid;

hybridizing the second target nucleic acid to the second set of capture extenders;

associating the second set of capture extenders with the solid support,

whereby hybridizing the second target nucleic acid to the second set of capture extenders and associating the second set of capture extenders with the solid support captures the second target nucleic acid on the solid support; and

detecting the presence of the second target nucleic acid on the solid support.

16. The method of claim 15, wherein the solid support is a substantially planar solid support, and wherein the first target nucleic acid is captured at a first selected position on the solid support and the second target nucleic acid is captured at a second selected position on the solid support.

17. The method of claim 15, wherein the solid support comprises a population of particles, the population comprising at least two sets of particles, the particles in each set being distinguishable from the particles in every other set;

wherein the first target nucleic acid is captured on a first set of the particles; and

wherein the second target nucleic acid is captured on a second set of the particles.

18. The method of claim 17, wherein the first set of particles comprises a first capture probe, which first capture probe is capable of hybridizing to the capture extenders comprising the first set of capture extenders;

and wherein the second set of particles comprises a second capture probe, which second capture probe is capable of hybridizing to the capture extenders comprising the second set of capture extenders.

19. The method of claim 1, wherein detecting the presence of the first target nucleic acid on the solid support comprises hybridizing a first set of one or more label extenders and a label probe system comprising a label to the first target nucleic acid and detecting the presence of the label on the solid support.

20. The method of claim 19, wherein the label probe system comprises an amplification multimer and a plurality of label probes, wherein the amplification multimer is capable of hybridizing simultaneously to a label extender and to a plurality of label probes.

21. The method of claim 20, wherein the label probe comprises the label.

22. The method of claim 19, wherein the label is a fluorescent label, and wherein detecting the presence of the label on the solid support comprises detecting a fluorescent signal from the label.

23. The method of claim 19, wherein the label is an enzyme.

24. The method of claim 1, wherein detecting the presence of the first target nucleic acid on the solid support comprises detecting an amount of the first target nucleic acid on the solid support.

25. The method of claim 1, comprising separating materials not captured on the solid support from the solid support.

26. A composition comprising:

a first set of n capture extenders, wherein n is at least two, which first set of capture extenders is capable of hybridizing to a first target nucleic acid, and which first set of capture extenders is associated with or is capable of being associated with a solid support; and

peripheral blood cell nucleic acids.

27. The composition of claim 26, comprising a whole blood lysate, which whole blood lysate comprises the peripheral blood cell nucleic acids.

28. The composition of claim 26, comprising the first target nucleic acid.

29. The composition of claim 26, wherein the peripheral blood cell nucleic acids comprise the first target nucleic acid.

30. The composition of claim 26, wherein the first target nucleic acid is an RNA.

31. The composition of claim 26, wherein the first target nucleic acid is a DNA.

32. The composition of claim 26, comprising an exogenously supplied protease.

33. The composition of claim 26, wherein n is at least three.

34. The composition of claim 26, wherein n is that most 10.

35. The composition of claim 26, comprising the solid support.

36. The composition of claim 35, wherein a first capture probe is bound to the solid support, the first capture probe being capable of hybridizing to the capture extenders of the first set of capture extenders and thereby associating the capture extenders with the solid support.

37. The composition of claim 35, wherein the solid support is a substantially planar solid support.

38. The composition of claim 35, wherein the solid support comprises a plurality of particles.

39. The composition of claim 26, comprising a second set of m capture extenders, wherein m is at least two, which second set of capture extenders is capable of hybridizing to a second target nucleic acid, and which second set of capture extenders is associated with or is capable of being associated with the solid support.

40. The composition of claim 39, comprising the second target nucleic acid.

41. The composition of claim 39, wherein the solid support is a substantially planar solid support, wherein the first set of capture extenders is associated with or is capable of being associated with a first selected position on the solid support, and wherein the second set of capture extenders is associated with or is capable of being associated with a second selected position on the solid support.

42. The composition of claim 39, wherein the solid support comprises a population of particles, the population comprising at least two sets of particles, the particles in each set being distinguishable from the particles in every other set;

wherein the first set of capture extenders is associated with or is capable of being associated with a first set of the particles; and

wherein the second set of capture extenders is associated with or is capable of being associated with a second set of the particles.

43. The composition of claim 42, wherein the first set of particles comprises a first capture probe, which first capture probe is capable of hybridizing to the capture extenders comprising the first set of capture extenders;

and wherein the second set of particles comprises a second capture probe, which second capture probe is capable of hybridizing to the capture extenders comprising the second set of capture extenders.

44. The composition of claim 26, comprising a first set of one or more label extenders, which first set of label extenders is capable of hybridizing to the first target nucleic acid, and/or a label probe system comprising a label.

45. The composition of claim 44, wherein the label probe system comprises an amplification multimer and a plurality of label probes, wherein the amplification multimer is capable of hybridizing simultaneously to a label extender and to a plurality of label probes.

46. The composition of claim 45, wherein the label probe comprises the label.

47. The composition of claim 44, wherein the label is a fluorescent label or an enzyme.

48. A method of detecting at least a first target nucleic acid, the method comprising:

providing plasma, which plasma comprises the first target nucleic acid;

contacting the plasma with a first set of n capture extenders, wherein n is at least two, which first set of capture extenders is capable of hybridizing to the first target nucleic acid;

hybridizing the first target nucleic acid to the first set of capture extenders;

associating the first set of capture extenders with a solid support,

whereby hybridizing the first target nucleic acid to the first set of capture extenders and associating the first set of capture extenders with the solid support captures the first target nucleic acid on the solid support; and

detecting the presence of the first target nucleic acid on the solid support.

49. The method of claim 48, wherein the first target nucleic acid is an RNA.

50. The method of claim 48, wherein the first target nucleic acid is a DNA.

51. The method of claim 48, comprising contacting the plasma with an exogenously supplied protease prior to contacting the plasma with the first set of capture extenders.

52. The method of claim 48, wherein a first capture probe is bound to the solid support, and wherein associating the first set of capture extenders with the solid support comprises hybridizing the capture extenders to the first capture probe.

53. The method of claim 48, wherein the solid support is a substantially planar solid support.

54. The method of claim 48, wherein the solid support comprises a plurality of particles.

55. The method of claim 48, wherein the plasma comprises a second target nucleic acid, the method comprising:

contacting the plasma with a second set of m capture extenders, wherein m is at least two, which second set of capture extenders is capable of hybridizing to the second target nucleic acid;

hybridizing the second target nucleic acid to the second set of capture extenders;

associating the second set of capture extenders with the solid support,

whereby hybridizing the second target nucleic acid to the second set of capture extenders and associating the second set of capture extenders with the solid support captures the second target nucleic acid on the solid support; and

detecting the presence of the second target nucleic acid on the solid support.

56. The method of claim 48, wherein detecting the presence of the first target nucleic acid on the solid support comprises hybridizing a first set of one or more label extenders and a label probe system comprising a label to the first target nucleic acid and detecting the presence of the label on the solid support.

57. A composition comprising:

a first set of n capture extenders, wherein n is at least two, which first set of capture extenders is capable of hybridizing to a first target nucleic acid, and which first set of capture extenders is associated with or is capable of being associated with a solid support; and

plasma.

58. The composition of claim 57, comprising the first target nucleic acid.

59. The composition of claim 57, wherein the first target nucleic acid is an RNA.

60. The composition of claim 57, wherein the first target nucleic acid is a DNA.

61. The composition of claim 57, comprising an exogenously supplied protease.

62. The composition of claim 57, comprising the solid support.

63. The composition of claim 62, wherein a first capture probe is bound to the solid support, the first capture probe being capable of hybridizing to the capture extenders of the first set of capture extenders and thereby associating the capture extenders with the solid support.

64. The composition of claim 57, comprising a second set of m capture extenders, wherein m is at least two, which second set of capture extenders is capable of hybridizing to a second target nucleic acid, and which second set of capture extenders is associated with or is capable of being associated with the solid support.

65. The composition of claim 64, wherein the solid support comprises a population of particles, the population comprising at least two sets of particles, the particles in each set being distinguishable from the particles in every other set;

wherein the first set of capture extenders is associated with or is capable of being associated with a first set of the particles; and

wherein the second set of capture extenders is associated with or is capable of being associated with a second set of the particles.

66. The composition of claim 65, wherein the first set of particles comprises a first capture probe, which first capture probe is capable of hybridizing to the capture extenders comprising the first set of capture extenders;

and wherein the second set of particles comprises a second capture probe, which second capture probe is capable of hybridizing to the capture extenders comprising the second set of capture extenders.

67. The composition of claim 57, comprising a first set of one or more label extenders, which first set of label extenders is capable of hybridizing to the first target nucleic acid, and/or a label probe system comprising a label.

68. A kit for detecting at least a first target nucleic acid, the kit comprising:

a first capture probe bound to a solid support;

a first set of n capture extenders, wherein n is at least two, which first set of capture extenders is capable of hybridizing to the first target nucleic acid and to the first capture probe;

a label probe system comprising a label;

a first set of one or more label extenders, which label extenders are capable of hybridizing to the first target nucleic acid and to the label probe system;

a first solution comprising a detergent;

a protease; and

instructions for detecting the first target nucleic acid in whole blood, in peripheral blood cells, and/or in plasma with the kit;

packaged in one or more containers.

69. The kit of claim 68, comprising:

a second capture probe bound to the solid support;

a second set of m capture extenders, wherein m is at least two, which second set of capture extenders is capable of hybridizing to a second target nucleic acid and to the second capture probe; and

a second set of one or more label extenders, which label extenders are capable of hybridizing to the second target nucleic acid and to the label probe system.