Method for examining reactivity and method for detecting a complex

• US 20040014124A1
• Filed: 06/30/2003
• Published: 01/22/2004
• Est. Priority Date: 08/31/2000
• Status: Abandoned Application

• Current Assignee: Canon Kabushiki Kaisha (Canon Inc.)
• Original Assignee: Canon Kabushiki Kaisha (Canon Inc.)
• Inventors: Suzuki, Tomohiro, Okamoto, Tadashi, Shimizu, Satoshi, Yamamoto, Nobuko
• Application Number:
  US10/608,804
• Publication Number:
  US 20040014124A1
• Time in Patent Office: Days
• US Class Current: 435/6
• CPC Class Codes: C12Q 1/6837 using probe arrays or probe chips C12Q1/6874 takes precedence
• CPC Class Codes: C12Q 2565/518 characterised by the immobilisation of the nucleic acid sample or target

Abstract

Multiple specimens, typically biological samples having different properties and origins, are bound onto matrix substrates, and oligonucleotides, proteins and drugs are spotted on each matrix in an array to examine those specimens at a time for multiple items.

Claims

45. A method of detecting a complex formed between an oligonucleotide of which base sequence is known and a component having a capability of binding to said oligonucleotide, comprising the steps of:

preparing at least one oligonucleotide of which base sequence is known;

preparing at least two liquid test samples potentially containing a component having a capability of binding to said oligonucleotide;

binding said oligonucleotide as a probe to a predetermined region on a solid substrate to produce a detection substrate;

arranging a plurality of spots of said test samples at a predetermined amount to form an array of said test samples within said region with said oligonucleotide bound thereto;

detecting whether a complex between said oligonucleotide and said component is present or not for each of said plurality of spots; and

determining whether or not said component is contained in each of said liquid test samples, or how strong its binding capability to said oligonucleotide is, based on said detection.

46. The detection method according to claim 45, wherein said oligonucleotide bound to said detection substrate has a base sequence with a base length of 2 to 100.

47. The detection method according to claim 45, wherein said liquid test samples are solutions each containing at least one nucleic acid of which base sequence is unknown, detection is made whether a complex between said oligonucleotide and said nucleic acid is formed or not for each of said test samples to thereby determine whether or not said nucleic acid contains a base sequence complementary to the known base sequence of said oligonucleotide functioning as said component having a capability of binding to said oligonucleotide.

48. The detection method according to claim 47, wherein said nucleic acid contained in each of said liquid test samples includes a set of mRNAs extracted from an organic tissue.

49. The detection method according to claim 47, wherein said nucleic acid contained in each of said liquid test samples includes a cDNA library prepared based on a set of mRNAs extracted from an organic tissue.

50. The detection method according to claim 47, wherein said nucleic acid contained in each of said liquid test samples has a base length of 2 to 5000.

51. The detection method according to claim 45, wherein said liquid test samples are solutions each containing at lest one protein, the proteins contained in said test samples being different from one another.

52. The detection method according to claim 45, wherein said liquid test samples are solutions each containing at lest one chemical, the chemicals contained in said test samples being different from one another.

53. The detection method according to claim 45, wherein said liquid test samples are extracts from different biological species, tissues or cells.

54. The detection method according to claim 45, wherein a plurality of oligonucleotides having known base sequences different from one another are used as a probe, and said detection substrate has a plurality of predetermined sections arranged in a matrix form to which said oligonucleotides are to be bound, respectively.

55. The detection method according to claim 54, wherein said plurality of oligonucleotides are bound to said sections to constitute a matrix at a density of 400/cm² or lower, said sections having the same area as one another.

56. The detection method according to claim 54, wherein said test samples are spotted in an array form in each of said sections to which said plurality of oligonucleotides having known base sequences different from one another are bound so that the spot positions in each section are arranged in the same way as one another.

57. The detection method according to claim 45, wherein said solid substrate used as said detection substrate is made of glass.

58. The detection method according to claim 45, wherein said oligonucleotide is fixed on the detection substrate by covalent bonds.

59. The detection method according to claim 58, wherein said oligonucleotide is fixed on the detection substrate by covalent bonds formed through a chemical reaction of maleimide groups introduced to a glass surface of the substrate used as said solid substrate with thiol (—

SH) groups possessed by said oligonucleotide.

60. The detection method according to claim 59, wherein said maleimide groups introduced to the glass surface is formed by first introducing amino groups to the glass surface and then reacting N-(6-maleimidocaproyloxy)succinimide with the amino groups.

61. The detection method according to claim 59, wherein said maleimide groups introduced to the glass surface is formed by first introducing amino groups to the glass surface and then reacting succinimidyl 4-(maleimidophenyl)butyrate with the amino groups.

62. The detection method according to claim 58, wherein said oligonucleotide is fixed on the detection substrate by covalent bonds through a chemical reaction of epoxy groups introduced to a glass surface of the substrate used as said solid substrate with amino groups possessed by said oligonucleotide.

63. The detection method according to claim 45, wherein said detection substrate has a surface previously partitioned to form a plurality of sections, and two or more different types of oligonucleotides of which base sequences are known are previously bound to the sections, respectively, in a matrix form.

66. The detection method according to claim 45, wherein each of the spots of said two or more test samples formed in each section has a diameter of 200 μ

m or lower.

67. The detection method according to claim 45, wherein the spots of said two or more test samples formed in each section is arranged at a density of 400/cm² or smaller.

68. The detection method according to claim 45, wherein each of the spots of said two or more test samples is formed by supplying a predetermined amount of a solution of said test samples by an ink-jet method.

69. The detection method according to claim 68, wherein said two or more test samples are spotted by an ink-jet method as a solution containing a nucleic acid with base length of 100 to 5000, respectively.

70. The detection method according to claim 69, wherein said two or more test samples are spotted by an ink-jet method as a solution containing a nucleic acid as a total concentrations of 0.05 to 500 μ

M, respectively.

71. The detection method according to claim 69, wherein said ink-jet method used for spotting is a bubble jet method.

72. The detection method according to claim 45, wherein the spots of test samples are formed by contacting a pin having a tip for collecting sample solutions with a solution of each test sample to allow the sample solution to adhere to the tip of said pin for taking a predetermined amount of the solution and then physically contacting the tip of said pin with a surface of the substrate to transfer said predetermined amount of the solution to the substrate surface.

73. The detection method according to claim 45, wherein the spots of test samples are formed by sucking a solution of each test sample using a capillary having a tip for sucking sample solutions thereinto and then physically contacting the tip of said capillary with a surface of the substrate to transfer a predetermined amount of the solution to the substrate surface.

64-1. The detection method according to claim 64, wherein said wall member has a thickness in the range of 1 to 20 μ

m.