Bioassay system including optical detection apparatuses, and method for detecting biomolecules

• US 7,767,441 B2
• Filed: 10/21/2008
• Issued: 08/03/2010
• Est. Priority Date: 10/25/2007
• Status: Active Grant

• Current Assignee: Personal Genomics Taiwan, Inc.
• Original Assignee: Industrial Technology Research Institute
• Inventors: Chu, Cheng-Wei, Pan, Chao-Chi, Yao, Bin-Cheng, Chang, Shang-Chia, Li, Yu-Tang, Chiou, Chung-Fan
• Primary Examiner(s): Lam; Ann Y
• Application Number:
  US12/255,044
• Publication Number:
  US 20090146076A1
• Time in Patent Office: 651 Days
• Field of Search: None
• US Class Current: 435/287.2
• CPC Class Codes: C12Q 1/6804 Nucleic acid analysis using immunogens immunoassay G01N33/53
• CPC Class Codes: C12Q 1/6869 Methods for sequencing
• CPC Class Codes: C12Q 1/6874 involving nucleic acid arrays, e.g. sequencing by hybridisation
• CPC Class Codes: C12Q 2535/101 Sanger sequencing method, i.e. oligonucleotide sequencing using primer elongation and dideoxynucleotides as chain terminators
• CPC Class Codes: C12Q 2565/518 characterised by the immobilisation of the nucleic acid sample or target
• CPC Class Codes: C12Q 2565/607 being a sensor, e.g. electrode
• CPC Class Codes: G01N 21/6454 using an integrated detector array

Abstract

A bioassay system is disclosed. The bioassay system may include a plurality of optical detection apparatuses, each of which includes a substrate having a light detector, and a linker site formed over the light detector, the linker site being treated to affix the biomolecule to the linker site. The linker site is proximate to the light detector and is spaced apart from the light detector by a distance of less than or equal to 100 micrometers. The light detector collects light emitted from the biomolecule within a solid angle of greater than or equal to 0.8 SI steridian. The optical detection apparatus may further include an excitation light source formed over the substrate so as to provide a light source for exciting a fluorophore attached to the biomolecule.

Claims

1. An apparatus for identifying a single biomolecule, comprising:

a substrate having a light detector; and

a linker site formed over the light detector, the linker site being treated to affix the biomolecule to the linker site;

wherein the linker site is proximate to the light detector and is spaced apart from the light detector by a distance of less than or equal to 100 micrometers.

2. The apparatus of claim 1, further comprising a blind sheet formed over the substrate, the blind sheet including a pinhole having a diameter, wherein the linker site is formed proximate to the pinhole.

3. The apparatus of claim 2, wherein the pinhole has a diameter of less than or equal to 1,000 nanometers.

4. The apparatus of claim 2, wherein the pinhole has a diameter of less than or equal to 200 nanometers.

5. The apparatus of claim 2, further comprising a filter layer formed between the substrate and the blind sheet.

6. The apparatus of claim 2, further comprising a microlens formed between the substrate and the blind sheet.

7. The apparatus of claim 1, wherein the distance is less than or equal to 25 micrometers.

8. The apparatus of claim 1, wherein the distance is less than or equal to 6 micrometers.

9. The apparatus of claim 1, wherein the light detector collects light from the biomolecule within a solid angle, the solid angle being greater than or equal to 0.8 SI steridian.

10. An optical detection system, comprising at least 10,000 apparatuses as recited in claim 1.

11. A method of sequencing a plurality of nucleic acid molecules, the method comprising the steps of:

affixing a plurality of nucleic acid molecules to the linker sites of the optical detection system of claim 10; and

performing nucleic acid sequencing of the nucleic acid molecules in parallel on the optical detection system.

12. A method of detecting a plurality of biomolecules, the method comprising the steps of:

affixing a plurality of biomolecules to the linker sites of the optical detection system of claim 10; and

detecting the biomolecules on the optical detection system in parallel.

13. An optical detection system, comprising at least 250,000 apparatuses as recited in claim 1.

14. An optical detection system, comprising at least 2,000,000 apparatuses as recited in claim 1.

15. An optical detection system, comprising at least 10,000,000 apparatuses as recited in claim 1.

16. A method of sequencing a nucleic acid, comprising the steps of:

affixing one nucleic acid molecule to the linker site of the apparatus of claim 1; and

performing nucleic acid sequencing of the nucleic acid molecule on the apparatus.

17. The method of claim 16, wherein the nucleic acid is affixed to the linker site by binding to a polymerase molecule affixed to the linker site.

18. The method of claim 16, wherein the nucleic acid sequencing comprises the step of adding labeled nucleotides to the apparatus.

19. The method of claim 18, wherein the nucleotides are labeled fluorescently.

20. The method of claim 19, wherein the nucleotides are labeled fluorescently on their terminal phosphate.

21. The method of claim 16, wherein the nucleic acid sequencing is base extension sequencing and includes the step of adding blocked and labeled nucleotides to the apparatus.

22. The method of claim 21, wherein the nucleotides are labeled fluorescently.

23. The method of claim 22, wherein the nucleotides have identical fluorescent labels and are added sequentially.

24. The method of claim 22, wherein the nucleotides have distinct fluorescent labels and are added simultaneously.

25. The method of claim 16, wherein the nucleic acid sequencing is a ligase-based sequencing.

26. The method of claim 16, wherein the nucleic acid is amplified at the linker site before nucleic acid sequencing.

27. The method of claim 16, wherein the sequence of the nucleic acid is unknown.

28. The method of claim 16, wherein the nucleic acid is detected with a label excited by Fö

rster resonance energy transfer (FRET).

29. The method of claim 16, wherein the nucleic acid is detected with a label by time-resolved fluorescence technology.

30. A method of detecting a biomolecule, comprising the steps of:

affixing one or more biomolecule to the linker site of the apparatus of claim 1; and

detecting the biomolecule on the apparatus.

31. The method of claim 30, wherein the biomolecule comprises a label.

32. The method of claim 31, wherein the label is fluorescent.

33. The method of claim 32, wherein the biomolecule comprises a moiety chosen from a polypeptide, antibody, lipid, vitamin, low molecular weight organic molecule, and polysaccharide.

34. The method of claim 33, wherein the biomolecule is affixed to the linker site of the apparatus by a linking molecule.

35. The method of claim 34, wherein the linking molecule comprises a capture molecule.

36. The method of claim 35, wherein the capture molecule is a protein.

37. The method of claim 35, wherein the capture molecule is an antibody.

38. The method of claim 35, wherein the linking molecule comprises a nucleic acid tag.

39. The method of claim 38, further comprising the step of detecting the nucleic acid tag of the linking molecule on the apparatus.

40. The method of claim 39, wherein the nucleic acid tag is detected by nucleic acid sequencing.

41. The method of claim 39, wherein the nucleic acid tag is detected by hybridization to a nucleic acid probe.

42. The method of claim 41, wherein the nucleic acid probe is labeled fluorescently.

43. A method of providing biomolecule analysis service, comprising the steps of:

providing a sample comprising a biomolecule from a service requester to a service provider;

the service requester receiving analytical results from the service provider, wherein the results are produced using the apparatus of claim 1.

44. The method of claim 43, wherein the method is performed for remunerative consideration.

45. The method of claim 44, wherein the service requester and the service provider are mediated by a vendor.

46. The method of claim 43, wherein the analytical results are produced in another country.

47. The method of claim 43, wherein the analytical results are produced in a country other than the United States of America.

48. An apparatus for identifying a single biomolecule, comprising:

a substrate having a light detector;

a linker site formed over the light detector, the linker site being treated to affix the biomolecule to the linker site; and

an excitation light source formed over the substrate;

wherein the linker site is proximate to the light detector and is spaced apart from the light detector by a distance of less than or equal to 100 micrometers.

49. The apparatus of claim 48, wherein the excitation light source includes a light emitting layer, the light emitting layer emitting excitation light to the linker site along a horizontal direction parallel to a surface of the light detector.

50. The apparatus of claim 49, further comprising a filter layer formed between the substrate and the light emitting layer.

51. The apparatus of claim 48, wherein the excitation light source is chosen from a light emitting diode (LED), an organic light emitting diode (OLED), a polymer light emitting diode (PLED), and a laser diode (LD).

52. The apparatus of claim 48, wherein the excitation light source provides excitation light of a first wavelength range not overlapping with a second wavelength range of light emitted from the biomolecule.

53. An apparatus for identifying a single biomolecule, comprising:

a substrate having a light detector; and

a linker site formed over the light detector, the linker site being treated to affix the biomolecule to the linker site;

wherein the light detector collects light emitted from the biomolecule within a solid angle of greater than or equal to 0.8 SI steridian.

54. An apparatus for identifying a single biomolecule, comprising:

a substrate having a light detector;

a linker site formed over the light detector, the linker site being treated to affix the biomolecule to it; and

an excitation light source formed over the substrate;

wherein the light detector collects light emitted from the biomolecule within a solid angle of greater than or equal to 0.8 SI steridian.

55. A method for manufacturing an apparatus for identifying a single biomolecule, comprising:

forming a light detector and a control circuit on a substrate;

forming a blind sheet having a pinhole over the substrate; and

forming a linker site over the light detector and proximate to the pinhole, the linker site being treated to affix the biomolecule to the linker site, wherein the linker site is proximate to the light detector and is spaced apart from the light detector by a distance of less than or equal to 100 micrometers.

56. The method of claim 55, further comprising forming a filter layer between the substrate and the blind sheet.

57. The method of claim 56, wherein forming the blind sheet comprises:

forming an opaque layer on the filter layer;

forming a photoresist layer on the opaque layer;

patterning the photoresist layer to expose a portion of the opaque layer;

etching the opaque layer using the patterned photoresist layer as a mask until the filter layer is exposed; and

removing the photoresist layer.

58. The method of claim 57, wherein the opaque layer comprises metal.