Nanochannel arrays and their preparation and use for high throughput macromolecular analysis

• US 7,670,770 B2
• Filed: 07/25/2002
• Issued: 03/02/2010
• Est. Priority Date: 07/25/2001
• Status: Expired due to Term

• Current Assignee: The Trustees of Princeton University (Princeton University)
• Original Assignee: The Trustees of Princeton University (Princeton University)
• Inventors: Yu, Zhaoning, Cao, Han, Tegenfeldt, Jonas O., Chou, Stephen Y., Austin, Robert H.
• Primary Examiner(s): FORMAN, BETTY J
• Application Number:
  US10/484,293
• Publication Number:
  US 20040197843A1
• Time in Patent Office: 2,777 Days
• Field of Search: None
• US Class Current: 435/6.12
• CPC Class Codes: B01J 2219/00274 Sequential or parallel reactions; Apparatus and devices for combinatorial chemistry or for making arrays; Chemical library technology
• CPC Class Codes: B01L 2200/0663 Stretching or orienting elongated molecules or particles
• CPC Class Codes: B01L 2200/12 Specific details about manufacturing devices
• CPC Class Codes: B01L 2300/0627 Sensor or part of a sensor is integrated
• CPC Class Codes: B01L 2300/0861 Configuration of multiple channels and/or chambers in a single devices
• CPC Class Codes: B01L 2300/0896 Nanoscaled
• CPC Class Codes: B01L 2400/0415 electrical forces, e.g. electrokinetic
• CPC Class Codes: B01L 2400/0418 electro-osmotic flow [EOF]
• CPC Class Codes: B01L 2400/086 using baffles or other fixed flow obstructions
• CPC Class Codes: B01L 3/502707 characterised by the manufacture of the container or its components by shaping or joining plastic parts B29C59/00, B29C65/00; by laminating B32B37/00; manufacture of microstructural devices in general B81C
• CPC Class Codes: B81B 1/002 Holes characterised by their shape, in either longitudinal or sectional plane
• CPC Class Codes: B81B 2201/058 Microfluidics not provided for in B81B2201/051 - B81B2201/054
• CPC Class Codes: B81B 2203/0338 Channels
• CPC Class Codes: B81C 1/00071 Channels
• CPC Class Codes: B82Y 30/00 Nanotechnology for materials or surface science, e.g. nanocomposites
• CPC Class Codes: C12Q 1/6837 using probe arrays or probe chips C12Q1/6874 takes precedence
• CPC Class Codes: C12Q 1/6869 Methods for sequencing
• CPC Class Codes: G01N 21/6428 Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes" in vivo A61B5/00; immunoassay G01N33/53
• CPC Class Codes: G01N 21/648 using evanescent coupling or surface plasmon coupling for the excitation of fluorescence
• CPC Class Codes: G01N 27/447 using electrophoresis
• CPC Class Codes: G01N 27/44791 : Microapparatus sample containers with integrated microfluidic structures B01L3/5027
• CPC Class Codes: G01N 33/48721 : Investigating individual macromolecules, e.g. by translocation through nanopores Coulter counters in general G01N15/12; fabrication methods for nanoscale apertures B81B1/00; sequencing of nucleic acids C12Q1/68
• CPC Class Codes: Y10S 977/704 : Nucleic acids, e.g. DNA or RNA
• CPC Class Codes: Y10S 977/88 : with arrangement, process, or apparatus for testing
• CPC Class Codes: Y10S 977/883 : Fluidic self-assembly, FSA

Abstract

Nanochannel arrays that enable high-throughput macromolecular analysis are disclosed. Also disclosed are methods of preparing nanochannel arrays and nanofluidic chips. Methods of analyzing macromolecules, such as entire strands of genomic DNA, are also disclosed, as well as systems for carrying out these methods.

Claims

1. A method of isolating, imaging, and analyzing, in parallel, two or more individual nucleic acid biopolymers, comprising the steps of:

providing a nanofluidic chip, comprising;

a) nanochannel array, comprising;

a surface having a plurality of parallel nanochannels running along the material of the surface, said nanochannels having a trench width of less than about 150 nanometers and a trench depth of less than 200 nanometers;

at least one of the plurality of nanochannels being surmounted by sealing material to render such nanochannels at least substantially enclosed;

at least two of the nanochannels capable of admitting a fluid;

b) at least one sample reservoir in fluid communication with at least two of the nanochannels, said sample reservoir capable of releasing a fluid containing at least two nucleic acid biopolymer;

providing the at least one sample reservoir with at least one fluid, said fluid comprising at least two nucleic acid biopolymers;

transporting the at least two nucleic acid biopolymers into the at least two nanochannels to elongate said at least two nucleic acid biopolymers, the at least two nucleic acid biopolymers being individually confined within the at least two nanochannels such that the at least two nucleic acid biopolymer are transported through the at least two nanochannels in an unfolded fashion;

imaging in parallel at least one signal transmitted from the at least two elongated and isolated nucleic acid biopolymers within the nanochannels; and

correlating the signal to at least one property of the at least two nucleic acid biopolymers to thereby analyze the two or more individual nucleic acid biopolymers.

2. The method according to claim 1 wherein the signal is correlated to at least one of the following properties:

length, conformation, and chemical composition.

3. The method of claim 1, wherein the two or more nucleic acid biopolymers are DNA and the signal is correlated to the base pair sequence of said DNA.

4. The method of claim 1 wherein a plurality of reservoirs provide a plurality of nucleic acid biopolymers into a plurality of nanochannels for determining the lengths of the macromolecules.

5. The method of claim 4 wherein more than two of the two or more nucleic acid biopolymers enter a single nanochannel.

6. The method of claim 1, wherein the two or more nucleic acid biopolymers are DNA or RNA.

7. The method of claim 1, wherein the two or more nucleic acid biopolymers are at least substantially unfolded in the channels.

8. The method of claim 1 wherein the concentration of the two or more nucleic acid biopolymers in the fluid is at least one attogram per milliliter.

9. The method of claim 1 wherein the concentration of the two or more nucleic acid biopolymers in the fluid is at least one femtogram per milliliter.

10. The method of claim 1 wherein the concentration of the two or more nucleic acid biopolymers in the fluid is at least one picogram per milliliter.

11. The method of claim 1 wherein the concentration of the two or more nucleic acid biopolymers in the fluid is less than 5 micrograms per milliliter.

12. The method of claim 1 wherein the concentration of the two or more nucleic acid biopolymers in the fluid is less than 0.5 micrograms per milliliter.

13. The method of claim 1 wherein the at least two nucleic acid biopolymers have an elongated length in the channels of greater than 150 nanometers.

14. The method of claim 1 wherein the at least two nucleic acid biopolymers have an elongated length in the channels of greater than 500 nanometers.

15. The method of claim 1 wherein the at least two nucleic acid biopolymers have an elongated length in the channels of greater than 1 micron.

16. The method of claim 1 wherein the at least two nucleic acid biopolymers have an elongated length in the channels of greater than 10 microns.

17. The method of claim 1 wherein the at least two nucleic acid biopolymers are DNA having greater than 10 base pairs.

18. The method of claim 1 wherein the at least two nucleic acid biopolymers are DNA having greater than 100 base pairs.

19. The method of claim 1 wherein the at least two nucleic acid biopolymers are DNA having greater than 1,000 base pairs.

20. The method of claim 1 wherein the at least two nucleic acid biopolymers are DNA having greater than 10,000 base pairs.

21. The method of claim 1 wherein the at least two nucleic acid biopolymers are DNA having greater than 20,000 base pairs.

22. The method of claim 1 wherein the at least two nucleic acid biopolymers are DNA having greater than 40,000 base pairs.

23. The method of claim 1 wherein the at least two nucleic acid biopolymers are DNA having greater than 80,000 base pairs.

24. The method of claim 1 wherein the nanochannel array has at least 96 sample reservoirs for simultaneously analyzing at least 96 different nucleic acid biopolymer fluid samples.

25. The method of claim 1 wherein at least one of the at least two nucleic acid biopolymers are restriction fragment length polymorphisms.

26. The method of claim 1 wherein least one of the at least two nucleic acid biopolymers is a chromosome.

27. The method of claim 26 wherein at least one chromosome is analyzed to determine the presence of at least one single nucleotide polymorphism.

28. The method of claim 1, further comprising the step of illuminating the at least two nucleic acid biopolymers within the at least two nanochannels.

29. The method of claim 1, wherein at least one of the at least two nanochannels is completely enclosed with sealing material on its surface.

30. The method of claim 1, wherein the nanochannel array comprises from more than 2 to about 500,000 parallel channels.

31. The method of claim 1, wherein the sealing material is optically transparent.

32. The method of claim 31, wherein at least one signal is transmitted through the optically transparent sealing material.

33. The method of claim 1, wherein one or more of the plurality of nanochannels is defined by walls that span the length of the substrate surface.