Compensator for multiple surface imaging

• US 9,068,220 B2
• Filed: 03/28/2014
• Issued: 06/30/2015
• Est. Priority Date: 05/05/2008
• Status: Active Grant

• Current Assignee: Illumina Incorporated
• Original Assignee: Illumina Incorporated
• Inventors: Feng, Wenyi, Bryant, Jason, Buermann, Dale
• Primary Examiner(s): Kim, Kiho
• Application Number:
  US14/229,455
• Publication Number:
  US 20140213486A1
• Time in Patent Office: 459 Days
• Field of Search: 250/458.1, 250/459.1
• US Class Current: 1/1
• CPC Class Codes: C12Q 1/6825 Nucleic acid detection involving sensors
• 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: G01N 2021/0346 Capillary cells; Microcells
• CPC Class Codes: G01N 2021/6419 Excitation at two or more wavelengths
• CPC Class Codes: G01N 2021/6421 Measuring at two or more wavelengths
• CPC Class Codes: G01N 21/05 Flow-through cuvettes G01N21/09 takes precedence; handling fluid samples G01N1/10
• CPC Class Codes: G01N 21/6458 Fluorescence microscopy fluorescence microscopes per se G02B21/0076 and G02B21/16
• CPC Class Codes: G02B 21/0024 Confocal scanning microscopes (CSOMs) or confocal "macroscopes"; Accessories which are not restricted to use with CSOMs, e.g. sample holders
• CPC Class Codes: G02B 2207/113 Fluorescence

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.

Claims

1. A system, comprising:

a flow cell comprising a first interior surface and a second interior surface in a generally mutually facing relation, wherein a plurality of nucleic acid sites is distributed on the first interior surface and on the second interior surface, and wherein the first interior surface and the second interior surface are separated by a fluid passage through which a fluorescent reagent flows to add fluorescent tags to the nucleic acid sites; and

a detection system configured to detect the fluorescent tags at a resolution to distinguish the nucleic acid sites on the first interior surface, and to detect fluorescent tags at a resolution to distinguish the nucleic acid sites on the second interior surface.

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

3. The system of claim 1, wherein the plurality of nucleic acid sites is distributed in a random spatial distribution on the first interior surface and on the second interior 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 interior surface and at successive regions of the second interior 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 interior surface and the second interior surface are detected from the same side of the flow cell.

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

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

12. The system of claim 9, wherein the first interior surface is disposed between a detector and the second interior 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 interior 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 interior surface is detected through a first objective and the second interior surface is detected through a second objective.

16. The system of claim 1, wherein the first interior surface and the second interior 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 interior surface and the second interior 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 interior 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 if 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.