Methods for concurrently processing multiple biological chip assays

• US 5,874,219 A
• Filed: 04/09/1996
• Issued: 02/23/1999
• Est. Priority Date: 06/07/1995
• Status: Expired due to Term

• Current Assignee: Affymetrix, Inc. (Thermo Fisher Scientific Incorporated)
• Original Assignee: Affymetrix, Inc. (Thermo Fisher Scientific Incorporated)
• Inventors: Fodor, Stephen P.A., Rava, Richard P., Trulson, Mark
• Primary Examiner(s): Campbell, Eggerton A.
• Application Number:
  US08/630,051
• Time in Patent Office: 1,050 Days
• Field of Search: 435/6, 435/5, 435/91.1, 435/91.2, 435/7.1, 435/7.9, 536/24.37, 536/33
• US Class Current: 506/9
• CPC Class Codes: B01J 19/0046 Sequential or parallel reactions, e.g. for the synthesis of polypeptides or polynucleotides; Apparatus and devices for combinatorial chemistry or for making molecular arrays synthesis methods per se C40B50/00
• CPC Class Codes: B01J 2219/00315 Microtiter plates
• CPC Class Codes: B01J 2219/00317 Microwell devices, i.e. having large numbers of wells
• CPC Class Codes: B01J 2219/00432 Photolithographic masks
• CPC Class Codes: B01J 2219/00529 DNA chips
• CPC Class Codes: B01J 2219/00608 DNA chips
• CPC Class Codes: B01J 2219/0061 The surface being organic
• CPC Class Codes: B01J 2219/00612 the surface being inorganic
• CPC Class Codes: B01J 2219/00617 by chemical means
• CPC Class Codes: B01J 2219/00621 by physical means, e.g. trenches, raised areas
• CPC Class Codes: B01J 2219/00626 Covalent
• CPC Class Codes: B01J 2219/00637 by coating it with another layer
• CPC Class Codes: B01J 2219/00659 Two-dimensional arrays
• CPC Class Codes: B01J 2219/00662 Two-dimensional arrays within two-dimensional arrays
• CPC Class Codes: B01J 2219/00702 Processes involving means for analysing and characterising the products
• CPC Class Codes: B01J 2219/00707 separated from the reactor apparatus
• CPC Class Codes: B01J 2219/00722 Nucleotides
• CPC Class Codes: B01L 2300/0636 Integrated biosensor, microarrays
• CPC Class Codes: B01L 2300/0829 Multi-well plates; Microtitration plates
• CPC Class Codes: B01L 3/5085 for multiple samples, e.g. microtitration plates
• CPC Class Codes: C12Q 1/6837 : using probe arrays or probe chips C12Q1/6874 takes precedence
• CPC Class Codes: C40B 40/06 : Libraries containing nucleotides or polynucleotides, or derivatives thereof
• CPC Class Codes: C40B 60/14 : for creating libraries
• CPC Class Codes: G01N 21/64 : Fluorescence; Phosphorescence
• CPC Class Codes: G01N 35/028 : having reaction cells in the form of microtitration plates

Abstract

Methods for concurrently processing multiple biological chip assays by providing a biological chip plate comprising a plurality of test wells, each test well having a biological chip having a molecular probe array; introducing samples into the test wells; subjecting the biological chip plate to manipulation by a fluid handling device that automatically performs steps to carry out reactions between target molecules in the samples and probes; and subjecting the biological chip plate to a biological chip plate reader that interrogates the probe arrays to detect any reactions between target molecules and probes.

Claims

1. A method for concurrently processing multiple biological chip assays comprising the steps of:

(a) providing a biological chip plate comprising a contiguous substrate that comprises a contiguous surface and, attached to the contiguous surface, a plurality of probe arrays and, surrounding the probe arrays, material resistant to the flow of liquid thereby forming a plurality of test wells, each test well defining a space for the introduction of a sample;

(b) manipulating the biological chip plate with a fluid handling device that automatically performs steps to carry out reactions between target molecules in a test sample and probes in a plurality of the test wells; and

(c) interrogating the probe arrays of the biological chip plate with a biological chip plate reader to detect reactions between target molecules and probes in a plurality of the test wells to generate assay results.

2. The method of claim 1 further comprising the step of processing the results with a computer.

3. The method of claim 2 wherein the results are processed into an image with the intensity in each region of the image varying according to the binding affinity between targets and probes.

4. The method of claim 1 wherein the probes are DNA or RNA molecules.

5. The method of claim 4 wherein

(a) the reactions are hybridization of target molecules to probes;

and

(b) the results provide information regarding the nucleotide sequence of the target molecule.

6. The method of claim 5 wherein the assay is the detection or identification of a pathogenic organism.

7. The method of claim 6 wherein the pathogenic organism is HIV.

8. The method of claim 5 wherein the assay is the detection or identification of a human nucleic acid.

9. The method of claim 8 wherein the assay is the detection of a human gene variant.

10. The method of claim 8 wherein the human gene variant indicates the existence of, or predisposition to cystic fibrosis, diabetes, muscular dystrophy or cancer.

11. The method of claim 1 wherein the assay is the identification of a probe in a library that binds to a receptor and the reaction is the binding of a target molecule to a peptide probe.

12. The method of claim 1 wherein the reaction is the catalytic transformation of a probe molecule by a target catalyst.

13. The method of claim 1 further comprising controlling the temperature of the samples in the test wells and adding or removing fluids from the test wells at predetermined times.

14. The method of claim 1 wherein the target molecule is tagged with a fluorescent marker, a chemiluminescent marker, a light scattering marker or a radioactive marker.

15. The method of claim 14 wherein the marker is a fluorescent marker selected from the group consisting of fluorescein, rhodamine, or cyanine.

16. The method of claim 1 wherein the fluid handling device and the biological chip plate reader comprise

(a) a confocal detection device having a monochromatic or polychromatic light source;

(b) optics for directing an excitation light from the light source at the substrate;

(c) a temperature controller for controlling temperature of the substrate during a reaction; and

(d) a detector for detecting fluorescence emitted by a fluorescently labelled molecule in response to the excitation light.

17. The method of claim 1 wherein the manipulating step comprises robotically controlling pipets for adding or removing fluids from the test wells at predetermined times.

18. The method of claim 16 wherein the detector comprises a photomultiplier tube.

19. The method of claim 16 wherein the interrogating step comprises focusing the excitation light to a point on the plate and determining the region from which the fluorescence originated.

20. The method of claim 16 wherein the target is tagged with a fluorescent marker and the biological chip plate reader comprises a CCD array.

21. The method of claim 1 wherein the probe arrays each have at least about 2500 features.

22. The method of claim 1 wherein the probe arrays each have at least about 50,000 features.

23. The method of claim 1 wherein each test well comprises an individual probe array.

24. The method of claim 1 wherein said chip plate is made by the steps of mating a wafer to an array of open ended wells.

25. The method of claim 5, wherein the samples are from a plurality of patients.

26. The method of claim 25 further comprising the step of using information from the results to determine which of the patients exhibit a genetic disease or characteristic.

27. The method of claim 1 wherein the interrogating step comprises the steps of:

(a) directing excitation light through a bottom surface of said plate; and

(b) detecting where targets bound on said probe arrays.

28. The method of claim 1 wherein the probe arrays are produced by light-directed probe synthesis.

29. The method of claim 1 wherein the interrogating step comprises determining the on- or off-rates of binding between target molecules in a sample and probes.

70. The method of claim 28 wherein said arrays of probes are made by repeating the steps of:

(a) selectively directing light at said surface to remove photoremovable protecting groups; and

(b) selectively coupling monomers to said surface.

30. A system for concurrently processing multiple biological chip assays comprising:

(a) a biological chip plate comprising a contiguous substrate that comprises a contiguous surface and, attached to the contiguous surface, a plurality of probe arrays and, surrounding the probe arrays, material resistant to the flow of liquid thereby forming a plurality of test wells, each test well defining a space for the introduction of a sample;

(b) a fluid handling device that automatically performs steps to carry out reactions between target molecules in the samples and probes in a plurality of the test wells; and

(c) a biological chip plate reader that interrogates the probe arrays to detect any reactions between target molecules and probes in a plurality of the test wells to produce assay results.

31. The system of claim 30 further comprising a computer comprising a program for processing the assay results.

32. The system of claim 30 wherein the probes are DNA or RNA molecules.

33. The system of claim 30 wherein the fluid handling device and the biological chip plate reader comprise

(a) a confocal detection device having a monochromatic or polychromatic light source;

(b) means for directing an excitation light from the light. source at the substrate;

(c) means for controlling temperature of the substrate during a reaction; and

(d) means for detecting fluorescence emitted by fluorescently labelled molecules in response to the excitation light.

34. The system of claim 33 wherein the fluid handling device comprises robotically controlled pipets for adding or removing fluids from the test wells at predetermined times;

and wherein

(a) the means for detecting the fluorescent emissions from the substrate comprise a photomultiplier tube;

(b) the means for focusing the excitation light to a point on the substrate and determining the region from which the fluorescence originated comprise an x-y-z translation table; and

(c) wherein translation of the x-y-z translation table, temperature control and data collection are managed and recorded by a digital computer.

35. The system of claim 33 wherein the means for directing excitation light and means for detecting are arranged to illuminate and read the test wells through a bottom side of the test wells.

36. The system of claim 30 wherein the biological chip plate reader comprises a line scanner.

37. The system of claim 30 wherein the biological chip plate reader comprises a CCD array.

38. The system of claim 30 wherein each probe array has at least about 2500 features.

39. The system of claim 30 wherein each probe array has at least about 50,000 features.

40. The system of claim 30 wherein the biological chip plate comprises about 96 test wells.

71. The system of claim 30 wherein the probe arrays are produced by light-directed probe synthesis.

41. A biological chip plate comprising a contiguous substrate that comprises on its surface a plurality of probe arrays and, surrounding the probe arrays, material resistant to the flow of liquid thereby forming a plurality of test wells, each test well defining a space for the introduction of a sample.

42. The biological chip plate of claim 41 wherein the substrate comprises functionalized glass or silica comprising Si, Ge, GaAs, GaP, SiO₂, SiN₄ or modified silicon.

43. The biological chip plate of claim 41 wherein the surface comprises polymers, plastics, resins, polysaccharides, silica or silica-based materials, carbon, metals, inorganic glasses, membranes, polymerized Langmuir Blodgett film, functionalized glass, (poly)tetrafluoroethylene, (poly)vinylidenedifluoride, polystyrene or polycarbonate.

44. The biological chip plate of claim 41 comprising a body having test wells, wherein each test well comprises an individual probe array.

45. The biological chip plate of claim 44 wherein the body comprises (poly)tetrafluoroethylene, (poly)vinylidenedifluoride, polypropylene, polystyrene, polycarbonate, or combinations thereof.

46. The biological chip plate of claim 41 comprising a wafer and a body, the wafer comprising a plurality of probe arrays and the body comprising a plurality of channels, wherein the body is attached to the surface of the wafer so that a plurality of the channels each cover a probe array and the wafer closes one end of a plurality of the channels, thereby forming the test wells.

47. The biological chip plate of claim 46 wherein the body comprises (poly)tetrafluoroethylene, (poly)vinylidenedifluoride, polypropylene, polystyrene, polycarbonate, or combinations thereof.

48. The biological chip plate of claim 41 comprising a wafer having a plurality of probe arrays and a material resistant to the flow of a liquid sample that surrounds each probe array.

49. The biological chip plate of claim 48 wherein the material is a hydrophobic material.

50. The biological chip plate of claim 41 comprising 96 wells arranged in 8 rows and 12 columns.

51. The biological chip plate of claim 41 wherein the probes are DNA or RNA molecules.

52. The biological chip plate of claim 51 wherein the DNA or RNA molecules comprise sequences directed to pathogenic organisms.

53. The biological chip plate of claim 52 wherein the pathogenic organism is HIV.

54. The biological chip plate of claim 51 wherein the DNA or RNA molecules comprise sequences directed to human DNA.

55. The biological chip plate of claim 52 wherein the sequences are directed to a human gene variant that indicates the existence of or predisposition to a genetic disease.

56. The biological chip plate of claim 55 wherein the disease is cystic fibrosis, diabetes, muscular dystrophy or cancer.

57. The biological chip plate of claim 41 wherein the probe arrays each of the test wells are the same.

58. The biological chip plate of claim 41 wherein the test wells are arranged in rows and columns and the rows contain the same probe array and the columns contain different probe arrays.

59. The biological chip plate of claim 41 wherein the probe arrays comprise at least 1000 features.

60. The biological chip plate of claim 41 wherein the probe arrays comprise at least 10,000 features.

61. The biological chip plate of claim 41 wherein the probe arrays comprise at least 100,000 features.

62. The biological chip plate of claim 41 wherein the probe arrays comprise at least 1,000,000 features.

72. The biological chip plate of claim 41 wherein the probe arrays are produced by light-directed probe synthesis.

63. A method for making a biological chip plate comprising the steps of providing a contiguous substrate and a body, the contiguous substrate that comprises a contiguous surface and, attached to the contiguous surface, a plurality of probe arrays, and the body comprising a plurality of channels;

and attaching the body to the surface of the contiguous substrate whereby a plurality of the channels each cover a probe array and the surface of the substrate closes one end of the plurality of channels, thereby forming test wells defining spaces for receiving samples.

64. The method of claim 63 wherein the probes are DNA or RNA molecules.

73. The method of claim 63 wherein the probe arrays are produced by light-directed probe synthesis.

65. A wafer comprising a contiguous substrate that comprises a contiguous surface and, attached to the contiguous surface, a plurality of probe arrays, wherein the probe arrays are arranged on the wafer surface in rows and columns, wherein the probe arrays in each row are the same and the probe arrays in each column are different.

66. The wafer of claim 65 wherein the probes are DNA or RNA molecules.

67. The wafer of claim 66 wherein the DNA or RNA molecules comprise sequences of HIV.

68. The wafer of claim 66 wherein the probe arrays in the columns comprise sequences for screening a plurality of genetic diseases.

69. The wafer of claim 68 wherein the genetic diseases comprise cystic fibrosis, diabetes, muscular dystrophy or cancer.

74. The wafer of claim 69 wherein the probe arrays are produced by light-directed probe synthesis.