Active-source-pixel, integrated device for rapid analysis of biological and chemical specimens

• US 10,712,273 B2
• Filed: 04/24/2018
• Issued: 07/14/2020
• Est. Priority Date: 11/17/2013
• Status: Active Grant

• Current Assignee: Quantum-Si Incorporated
• Original Assignee: Quantum-Si Incorporated
• Inventors: Rothberg, Jonathan M., Kabiri, Ali, Sickler, Jason W., Gyarfas, Brett J., Lackey, Jeremy, Schmid, Gerard
• Primary Examiner(s): Priest, Aaron A
• Application Number:
  US15/960,751
• Publication Number:
  US 20180348132A1
• Time in Patent Office: 812 Days
• Field of Search: None
• CPC Class Codes: B01L 2200/12 Specific details about manufacturing devices
• CPC Class Codes: B01L 2300/0829 Multi-well plates; Microtitration plates
• CPC Class Codes: B01L 2300/0887 Laminated structure
• CPC Class Codes: B01L 2300/0893 having a very large number of wells, microfabricated wells
• CPC Class Codes: B01L 2300/168 Specific optical properties, e.g. reflective coatings
• CPC Class Codes: B01L 3/5085 for multiple samples, e.g. microtitration plates
• 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 2521/101 DNA polymerase
• CPC Class Codes: C12Q 2525/101 incorporating non-naturally occurring nucleotides, e.g. inosine
• CPC Class Codes: C12Q 2537/157 A reaction step characterised by the number of molecules incorporated or released
• CPC Class Codes: C12Q 2563/107 fluorescence
• CPC Class Codes: C12Q 2565/607 being a sensor, e.g. electrode
• CPC Class Codes: G01N 2021/6419 Excitation at two or more wavelengths
• CPC Class Codes: G01N 2021/6441 with two or more labels
• CPC Class Codes: G01N 2021/6478 Special lenses
• CPC Class Codes: G01N 21/64 Fluorescence; Phosphorescence
• CPC Class Codes: G01N 21/645 Specially adapted constructive features of fluorimeters
• CPC Class Codes: G01N 21/6452 Individual samples arranged in a regular 2D-array, e.g. multiwell plates
• CPC Class Codes: G01N 21/6454 using an integrated detector array
• CPC Class Codes: G01N 21/648 : using evanescent coupling or surface plasmon coupling for the excitation of fluorescence
• CPC Class Codes: G01N 21/6486 : Measuring fluorescence of biological material, e.g. DNA, RNA, cells G01N21/6428 takes precedence
• CPC Class Codes: G01N 21/7743 : the reagent-coated grating coupling light in or out of the waveguide
• CPC Class Codes: G01N 21/7746 : the waveguide coupled to a cavity resonator
• CPC Class Codes: G01N 2201/02 : Mechanical
• CPC Class Codes: G01N 2201/06113 : Coherent sources; lasers
• CPC Class Codes: G01N 2201/0612 : Laser diodes
• CPC Class Codes: G01N 2201/062 : LED's
• CPC Class Codes: G01N 2201/068 : Optics, miscellaneous
• CPC Class Codes: G01N 2201/125 : Digital circuitry
• CPC Class Codes: Y10T 29/49016 : Antenna or wave energy "plumbing" making

Abstract

An active-source-pixel, integrated device capable of performing biomolecule detection and/or analysis, such as single-molecule nucleic acid sequencing, is described. An active pixel of the integrated device includes a sample well into which a sample to be analyzed may diffuse, an excitation source for providing excitation energy to the sample well, and a sensor configured to detect emission from the sample. The sensor may comprise two or more segments that produce a set of signals that are analyzed to differentiate between and identify tags that are attached to, or associated with, the sample. Tag differentiation may be spectral and/or temporal based. Identification of the tags may be used to detect, analyze, and/or sequence the biomolecule.

Claims

1. An integrated device for analyzing a plurality of samples in parallel, the device comprising:

a plurality of pixels arranged on a substrate, wherein an individual pixel of the plurality of pixels comprises;

a sample well having an excitation region configured to retain a biological sample;

a first structure located adjacent to or within the sample well and configured to affect coupling of at least excitation energy into the excitation region; and

a sensing system including a sensor that is configured to discriminate between at least two different emissions from the sample well, wherein the at least two different emissions comprise spectral and/or temporal differences;

at least one excitation source on the substrate that is arranged to provide the excitation energy to the sample well; and

circuitry arranged on the substrate to receive at least one signal from the sensor.

2. The integrated device of claim 1, wherein the first structure comprises a three-dimensional, micro- or nanoscale plasmonic structure that improves coupling of the excitation energy into the excitation region via plasmons excited at the first structure.

3. The integrated device of claim 1, wherein the first structure comprises a micro-optical or nano-optical structure that increases an intensity of the excitation energy within the excitation region.

4. The integrated device of claim 1, wherein the pixel further comprises a second structure located adjacent to the sample well that is configured to direct emitted radiations from the sample well into a plurality of different spatial distributions that are dependent upon wavelengths of the emitted radiations and to condense the emitted radiations in each of a plurality of different directions.

5. The integrated device of claim 1, wherein the plurality of pixels are arranged in an N×

M array on the substrate, where N is a number of pixels the array extends in a first direction and M is a number of pixels the array extends in a second direction, wherein each of N and M has a value between 50 pixels and 100000 pixels.

6. The integrated device of claim 1, further comprising a walled chamber formed around the plurality of pixels that is configured to hold a specimen.

7. The integrated device of claim 1, further comprising a divot extending from the sample well into an optically-transparent material, wherein the excitation region includes the divot.

8. The integrated device of claim 1, wherein the integrated device and sample well are arranged to receive samples in a fluid suspension.

9. The integrated device of claim 1, wherein the sample well comprises an entrance aperture at which excitation energy is received.

10. The integrated device of claim 9, wherein a diameter of the entrance aperture is between approximately 30 nanometers and approximately 250 nanometers.

11. The integrated device of claim 1, further comprising a reflector spaced from the sample well, wherein the reflector and material in which the sample well is formed comprises a resonant cavity that increases an intensity of excitation energy at the excitation region.

12. The integrated device of claim 1, further comprising an adherent arranged within the excitation region, wherein the adherent is configured to retain a sample in the excitation region.

13. The integrated device of claim 12, wherein the adherent comprises a biological or chemical substance formed on at least one surface within the sample well.

14. The integrated device of claim 1, wherein the first structure comprises a resonant structure for a characteristic frequency of the excitation energy and the sample well is positioned at a position corresponding to a coupling region of the resonant structure.

15. The integrated device of claim 14, wherein the resonant structure is a linear resonator or ring resonator.

16. The integrated device of claim 2, wherein the micro- or nanoscale plasmonic structure comprises a nano-antenna.

17. The integrated device of claim 2, wherein the micro- or nanoscale plasmonic structure comprises a resonant structure.

18. The integrated device of claim 2, wherein the plasmonic structure is configured to resonate at more than one frequency, wherein the more than one frequency includes a characteristic frequency of the excitation energy and/or characteristic frequencies of the emissions from the sample well.

19. The integrated device of claim 3, wherein the first structure comprises a diffractive optical element configured to concentrate the excitation energy at the excitation region.

20. The integrated device of claim 3, wherein the first structure comprises a thin lossy film adjacent the sample well.

21. The integrated device of claim 20, further comprising a dielectric layer disposed between the thin lossy film and the sample well.

22. The integrated device of claim 20, wherein an index of refraction of the thin lossy film is within a same order of magnitude as an extinction coefficient for the thin lossy film at a wavelength of the excitation energy.

23. The integrated device of claim 20, wherein the thin lossy film is formed of silicon or germanium, or a combination thereof.

24. The integrated device of claim 20, wherein a thickness of the thin lossy film is between approximately 3 nm and approximately 80 nm.

25. The integrated device of claim 20, wherein the thin lossy film includes multiple layers of at least two different materials.

26. The integrated device of claim 3, wherein the first structure comprises a microcavity located adjacent to the sample well and configured to concentrate the excitation energy at the excitation region.

27. The integrated device of claim 1, wherein the sensing system comprises a filter configured to suppress an amount of excitation energy at the sensor.

28. The integrated device of claim 27, wherein the filter comprises a multi-layer stack.

29. The integrated device of claim 27, wherein the filter comprises a frequency-selective surface.

30. The integrated device of claim 1, wherein the sensing system comprises an optical block that blocks at least some excitation energy from the excitation source.

31. The integrated device of claim 1, wherein the sensing system comprises at least one phase mask.

32. The integrated device of claim 4, wherein the second structure comprises a plasmonic structure, wherein the plasmonic structure is formed in an optical near field of the sample well.

33. The integrated device of claim 32, wherein the plasmonic structure is formed at least in part in a same layer in which the sample well is formed.

34. The integrated device of claim 32, wherein the plasmonic structure comprises a plurality of annular projections formed in a layer and centered on the sample well.

35. The integrated device of claim 32, wherein the plasmonic structure comprises a spiral grating.

36. The integrated device of claim 32, wherein the plasmonic structure comprises a nano-antenna array having an array of circular holes distributed around the sample well and wherein the circular holes are formed in a conductive layer.

37. The integrated device of claim 36, wherein the conductive layer in which the circular holes are formed is a same layer in which the sample well is formed.

38. The integrated device of claim 36, wherein the nano-antenna array comprises an array of circular discs distributed around the sample well, wherein the circular discs are formed in a conductive material.

39. The integrated device of claim 4, wherein the second structure comprises a circular grating, a spiral grating, or an aperture array formed in an optical far field of the sample well.

40. The integrated device of claim 1, wherein the sensor comprises a bullseye photodetector having a central light detector and at least one ring-shaped light detector spaced from and surrounding the central light detector.

41. The integrated device of claim 1, wherein the sensor comprises at least two spatially-separated detectors.

42. The integrated device of claim 40, further comprising read-out circuitry configured to discern four different emission energies from the sample detected by two or more spatially-separated detectors, wherein the sensor and read-out circuitry comprise CMOS circuit elements.

43. The integrated device of claim 1, wherein the at least one excitation source comprises at least one source of optical radiation having a characteristic wavelength between approximately 350 nm and approximately 1000 nm and is configured to excite at least one sample well.

44. The integrated device of claim 1, wherein the at least one excitation source is located between the sample well and the sensor.

45. The integrated device of claim 1, wherein the at least one excitation source comprises an organic light-emitting diode (OLED), wherein an electrode of the OLED is formed from a conductive layer in which the sample well is formed and further comprising surface features patterned in the electrode adjacent to the sample well and arranged to enhance a surface wave that is excited by the OLED and delivers excitation energy to the excitation region.

46. The integrated device of claim 1, wherein the circuitry comprises CMOS circuitry that includes multi-channel analog-to-digital converters.

47. The integrated device of claim 46, wherein the CMOS circuitry includes at least one field-programmable gate array (FPGA) and/or at least one application-specific integrated circuit (ASIC).

48. A portable instrument comprising:

at least one processor;

a dock configured to receive an integrated device having;

a plurality of pixels arranged on a substrate, wherein an individual pixel of the plurality of pixels comprises;

a sample well having an excitation region configured to retain a biological sample;

a first structure located adjacent to or within the sample well and configured to affect coupling of at least excitation energy into the excitation region; and

a sensing system including a sensor that is configured to discriminate between at least two different emissions from the sample well, wherein the at least two different emissions comprise spectral and/or temporal differences; and

circuitry arranged on the substrate to receive at least one signal from the sensor;

a cover configured to exclude a majority of external light from entering the dock; and

a first plurality of electrical contacts configured to connect to a second plurality of electrical contacts on the integrated device, wherein power may be provide to the integrated device through at least some of the first plurality of electrical contacts and the at least one signal from each sensor may be received through at least some of the first plurality of electrical contacts.

49. The portable instrument of claim 48, wherein the first plurality of electrical contacts is formed on a user-removable interposer.