Methods and apparatus for measuring analytes using large scale FET arrays

• US 9,194,000 B2
• Filed: 10/22/2014
• Issued: 11/24/2015
• Est. Priority Date: 06/25/2008
• Status: Active Grant

• Current Assignee: Life Technologies Corporation (Thermo Fisher Scientific Incorporated)
• Original Assignee: Life Technologies Corporation (Thermo Fisher Scientific Incorporated)
• Inventors: Rothberg, Jonathan M., Hinz, Wolfgang, Bustillo, James, Schultz, Jonathan, Johnson, Kim L., Leamon, John
• Primary Examiner(s): Ball, J. Christopher
• Application Number:
  US14/520,558
• Publication Number:
  US 20150126378A1
• Time in Patent Office: 398 Days
• Field of Search: None
• US Class Current: 1/1
• CPC Class Codes: B01L 2300/046 Function or devices integrated in the closure
• CPC Class Codes: B01L 2300/0663 Whole sensors
• CPC Class Codes: B01L 2300/0851 Bottom walls
• CPC Class Codes: B01L 2400/086 using baffles or other fixed flow obstructions
• CPC Class Codes: B01L 3/502761 specially adapted for handling suspended solids or molecules independently from the bulk fluid flow, e.g. for trapping or sorting beads, for physically stretching molecules investigating characteristics of particles G01N15/00
• 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 2565/301 Pyrophosphate (PPi)
• CPC Class Codes: C12Q 2565/607 being a sensor, e.g. electrode
• CPC Class Codes: C12Q 2565/629 being a microfluidic device
• CPC Class Codes: G01N 27/4145 specially adapted for biomolecules, e.g. gate electrode with immobilised receptors
• CPC Class Codes: G01N 27/4148 Integrated circuits therefor, e.g. fabricated by CMOS processing CMOS processing per se H01L21/82

Abstract

Methods and apparatus relating to very large scale FET arrays for analyte measurements. ChemFET (e.g., ISFET) arrays may be fabricated using conventional CMOS processing techniques based on improved FET pixel and array designs that increase measurement sensitivity and accuracy, and at the same time facilitate significantly small pixel sizes and dense arrays. Improved array control techniques provide for rapid data acquisition from large and dense arrays. Such arrays may be employed to detect a presence and/or concentration changes of various analyte types in a wide variety of chemical and/or biological processes. In one example, chemFET arrays facilitate DNA sequencing techniques based on monitoring changes in the concentration of inorganic pyrophosphate (PPi), hydrogen ions, and nucleotide triphosphates.

Claims

1. A method of making a bead array, the method comprising:

contacting a substrate having a surface comprising discrete reaction sites disposed within a flow cell with a solution introduced into the flow cell and including a population of different beads, the surface includes 10³ of the discrete reaction sites; and

applying centrifugal force to the substrate or the solution, or both, such that at least a subpopulation of the population of different beads associates with the discrete reaction sites.

2. The method of claim 1, wherein the surface of the substrate includes 10⁵ of the discrete reaction sites.

3. The method of claim 2, wherein the surface of the substrate includes 10⁶ of the discrete reaction sites.

4. The method of claim 1, wherein a center distance between adjacent reaction sites of the reaction sites is in a range of 1 micrometers to 9 micrometers.

5. The method of claim 1, wherein a ratio of the diameter of a reaction site of the discrete reaction sites to a diameter of a bead of the population of different beads is at least 0.7.

6. The method of claim 1, wherein a ratio of the height to the width or diameter of a reaction site of the discrete reaction sites is in a range of 1:

1 to 1.5;

1.

7. The method of claim 1, wherein a bead of the population of different beads has a size in a range of 1 micrometers to 10 micrometers.

8. The method of claim 1, wherein a bead of the population of different beads is bound to a template nucleic acid.

9. The method of claim 1, further comprising sequencing the template nucleic acid without optical detection.

10. A method of making a bead array, the method comprising:

contacting a substrate having a surface comprising microwells disposed within a flow cell with a solution introduced into the flow cell and including a population of beads conjugated to different nucleic acid templates; and

applying centrifugal force to the substrate and the solution to associate at least a subpopulation of beads of the population of beads with at least a subset of the microwells.

11. The method of claim 10, wherein the surface of the substrate includes 10⁵ of the microwells.

12. The method of claim 11, wherein the surface of the substrate includes 10⁶ of the microwells.

13. The method of claim 10, wherein a center distance between adjacent microwells is in a range of 1 micrometers to 9 micrometers.

14. The method of claim 10, wherein a ratio of the diameter of a microwell of the microwells to a diameter of a bead of the population of different beads is at least 0.7.

15. The method of claim 10, wherein a ratio of the height to the width or diameter of a microwell of the microwells is in a range of 1:

1 to 1.5;

1.

16. The method of claim 10, wherein a bead of the population of beads has a size in a range of 1 micrometers to 10 micrometers.

17. The method of claim 10, further comprising sequencing the different template nucleic acids without optical detection.

18. A method of making a bead array, the method comprising:

contacting a substrate having a surface comprising microwells disposed in a flow cell with a solution introduced into the flow cell and including a population of beads conjugated to different nucleic acid templates, a ratio of the diameter of a microwell of the microwells to a diameter of a bead of the population of different beads is at least 0.7, a ratio of the height to the width or diameter of a microwell of the microwells is in a range of 1;

1 to 1.5;

1; and

applying centrifugal force to the substrate and the solution to associate at least a subpopulation of beads of the population of beads with at least a subset of the microwells.

19. The method of claim 18, further comprising sequencing the different template nucleic acids without optical detection.

20. The method of claim 18, wherein a bead of the population of beads has a size in a range of 1 micrometers to 10 micrometers.