Compensator for multiple surface imaging

Patent metadata

Field	Value
Publication number	US-RE48561-E
Application number	US-201816000720-A
Country	US
Kind code	E1
Filing date	Jun 5, 2018
Priority date	May 5, 2008
Publication date	May 18, 2021
Grant date	May 18, 2021

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 method of manufacturing a biological evaluation support structure, the method comprising:
providing a first substrate having a first surface on which a first plurality of biological sample sites are disposed; providing a second substrate having a second surface on which a second plurality of biological sample sites are disposed; forming a patterned adhesive on at least one of the first surface of the first substrate or the second surface of the second substrate; and bonding the first substrate to the second substrate via the patterned adhesive, such that the second surface of the second substrate, with the second plurality of biological sample sites thereon, faces the first surface of the first substrate, with the first plurality of biological sample sites thereon, so as to form a biological evaluation support structure.

29. The method of claim 28, wherein the first plurality of biological sample sites are disposed in a predetermined pattern on the first surface of the first substrate, and the second plurality of biological sample sites are disposed in a predetermined pattern on the second surface of the second substrate.

30. The method of claim 29, wherein each of the biological sample sites of the first plurality of biological sample sites comprises a well in the first surface of the first substrate, and each of the biological sample sites of the second plurality of biological sample sites comprises a well in the second surface of the second substrate.

31. The method of claim 28, wherein the adhesive is patterned so as to form a plurality of flow passages through which reagent fluids are flowable.

32. The method of claim 28, wherein the adhesive comprises an epoxy resin.

33. The method of claim 28, wherein the patterned adhesive is printed on the at least one of the first substrate or the second substrate.

34. The method of claim 28, where the adhesive is a heat-curable adhesive, and the method further comprises curing the adhesive by heating the adhesive.

35. The method of claim 28, wherein the biological evaluation support structure is a flow cell.

36. The method of claim 28, wherein, in bonding the first substrate to the second substrate via the patterned adhesive, the first substrate is bonded to the second substrate via both the patterned adhesive and an intermediate layer.

37. The method of claim 36, wherein the intermediate layer is made of a polymer material.

38. The method of claim 36, wherein forming the patterned adhesive comprises forming a first patterned adhesive on the first substrate and forming a second patterned adhesive on the second substrate.

39. The method of claim 38, wherein forming the first patterned adhesive and forming the second patterned adhesive comprise printing the first patterned adhesive onto the first substrate and printing the second patterned adhesive onto the second substrate.

40. The method of claim 28, wherein the second substrate has a plurality of ports extending therethrough.

Assignees

• Illumina Inc

Inventors

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

Classifications

• G01N21/05 Primary

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

• G02B21/0024

  Confocal scanning microscopes (CSOMs) or confocal "macroscopes"; Accessories which are not restricted to use with CSOMs, e.g. sample holders · CPC title

• G02B2207/113

  Fluorescence · CPC title

• G01N21/6458

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

• G01N2021/0346

  Capillary cells; Microcells · CPC title