Compensator for multiple surface imaging

Patent metadata

Field	Value
Publication number	US-RE49884-E
Application number	US-202117214093-A
Country	US
Kind code	E1
Filing date	Jun 8, 2021
Priority date	May 5, 2008
Publication date	Mar 26, 2024
Grant date	Mar 26, 2024

Abstract

A system and method for imaging biological samples on multiple surfaces of a support structure are disclosed. The support structure may be a flow cell through which a reagent fluid is allowed to flow and interact with the biological samples. Excitation radiation from at least one radiation source may be used to excite the biological samples on multiple surfaces. In this manner, fluorescent emission radiation may be generated from the biological samples and subsequently captured and detected by detection optics and at least one detector. The detected fluorescent emission radiation may then be used to generate image data. This imaging of multiple surfaces may be accomplished either sequentially or simultaneously. In addition, the techniques of the present invention may be used with any type of imaging system. For instance, both epifluorescent and total internal reflection methods may benefit from the techniques of the present invention.

First claim

The invention claimed is:

1. A system, comprising:
a flow cell comprising first and second surfaces separated by a fluid passage through which a fluorescent reagent flows to add fluorescent tags to nucleic acid sites distributed on the first and second surfaces; and a detection system configured to detect the fluorescent tags to distinguish the nucleic acid sites on the first surface, and to detect the fluorescent tags to distinguish the nucleic acid sites on the second surface.

2. The system of claim 1 , wherein the nucleic acid sites are distributed in a spatially ordered pattern on the first surface and on the second surface.

3. The system of claim 1 , wherein the nucleic acid sites are distributed in a random spatial distribution on the first surface and on the second surface.

4. The system of claim 1 , wherein the nucleic acid sites are separated with spaces between each other.

5. The system of claim 1 , wherein the nucleic acid sites are present at a density of at least 1,000 sites per square millimeter.

6. The system of claim 1 , wherein the detection system is configured to detect the fluorescent tags at an optical resolution between 0.1 and 50 microns.

7. The system of claim 1 , wherein the flow cell is configured to be translated, and the detection system if configured to detect the fluorescent tags at successive regions of the first surface and at successive regions of the second surface, respectively.

8. The system of claim 7 , wherein the detection system is configured to perform wide angle area detection of each successive region.

9. The system of claim 1 , wherein the first surface and the second surface are detected from the same side of the flow cell.

10. The system of claim 9 , wherein the first surface is disposed between an excitation source and the second surface during detection by the detection system.

11. The system of claim 10 , wherein the first surface and the second surface are excited by total internal reflection.

12. The system of claim 9 , wherein the first surface is disposed between a detector and the second surface during detection by the detection system.

13. The system of claim 12 , comprising corrective optics configured to be inserted and removed between the detector and the flow cell after detection of the fluorescent tags on the first surface by the detection system.

14. The system of claim 13 , wherein the corrective optics comprise a lens, objective, or cover slip.

15. The system of claim 12 , wherein the first surface is detected through a first objective and the second surface is detected through a second objective.

16. The system of claim 1 , wherein the first surface and the second surface are detected from opposite sides of the flow cell.

17. The system of claim 1 , wherein the detection system is configured to produce one or more images of the first surface and the second surface.

18. The system of claim 17 , wherein the one or more images have a resolution of 10 microns or less.

19. The system of claim 1 , comprising a radiation source configured to direct excitation radiation toward the first and second surfaces at several different wavelengths.

20. The system of claim 19 , wherein the detection system is configured to capture and detect emitted radiation returned in response to each wavelength.

21. The system of claim 1 , wherein the detection system is configured to perform confocal detection of fluorescence emitted from the nucleic acid sites.

22. The system of claim 1 , wherein the detection system is configured to repeat detection of the fluorescent tags in a process of sequencing the nucleic acids at the nucleic acid sites.

23. The system of claim 22 , wherein the sequencing comprises sequencing by synthesis.

24. The system of claim 22 , wherein the sequencing comprises sequencing by ligation.

25. The system of claim 1 , wherein the fluorescent reagent comprises fluorescently labeled nucleic acids.

26. The system of claim 1 , wherein the fluorescent reagent comprises fluorescently labeled nucleotides.

27. The system of claim 1 , wherein each of the nucleic acid sites constitutes a population of nucleic acids having identical sequences.

28. A biological sample imaging system, comprising:
a support structure comprising a flow lane defined by a bottom surface and a top surface; a prism supporting the support structure; a radiation source configured to emit a radiation beam; a translation system configured to move the support structure and prism relative to the radiation source; and a detector configured to:
detect radiation from emissive components of first biological samples disposed on the bottom surface while the support structure and the prism are in a first position relative to the radiation source, and
detect radiation from emissive components of second biological samples disposed on the top surface while the support structure and the prism are in a second position relative to the radiation source.

29. The system of claim 28, wherein:
the first biological samples are distributed in a spatially ordered pattern on the bottom surface, and the second biological samples are distributed in a spatially ordered pattern on the top surface.

30. The system of claim 28, wherein:
the first biological samples are distributed in a random spatial distribution on the bottom surface, and the second biological samples are distributed in a random spatial distribution on the top surface.

31. The system of claim 28, wherein:
the first biological samples are separated with spaces between each other, and the second biological samples separated with spaces between each other.

32. The system of claim 28, wherein:
the first biological samples are present at a density of at least 1,000 sites per square millimeter, and the second biological samples are present at a density of at least 1,000 sites per square millimeter.

33. The system of claim 28, wherein the detector is configured to detect the emissive components of the first and second biological samples at an optical resolution between 0.1 and 50 microns.

34. The system of claim 28, wherein the detector is configured to detect the emissive components of the first biological samples disposed on the bottom surface and the emissive components of the second biological samples disposed on the top surface from the same side of the support structure.

35. The system of claim 28, wherein:
the radiation source is configured such that, while the support structure and the prism are in the first position relative to the radiation source, the radiation source emits the radiation beam through the prism toward the bottom surface defining the flow lane, such that an emission beam is emitted from the emissive components of the first biological samples toward the detector; and the radiation source is configured such that, while the support structure and the prism are in the second position relative to the radiation source, the radiation source emits the radiation beam through the prism and the support structure until the radiation beam contacts an air/glass interface of the support structure, at which point the radiation beam is redirected toward the top surface defining the flow lane, such that an emission beam is emitted from

Assignees

• Illumina Inc

Inventors

• Feng Wenyi
• Bryant Jason
• Barnard Steven
• Bacigalupo Maria Candelaria Rogert

Classifications

• G01N21/6458 Primary

  Fluorescence microscopy (fluorescence microscopes per se G02B21/0076 and G02B21/16) · CPC title

• C12Q1/6825

  Nucleic acid detection involving sensors · CPC title

• C12Q1/6869

  Methods for sequencing · CPC title

• C12Q1/6874 Primary

  involving nucleic acid arrays, e.g. sequencing by hybridisation · CPC title

• G01N21/05 Primary

  Flow-through cuvettes (G01N21/09 takes precedence; handling fluid samples G01N1/10) · CPC title