Integrated optoelectronic read head and fluidic cartridge useful for nucleic acid sequencing

• US 9,650,669 B2
• Filed: 07/18/2014
• Issued: 05/16/2017
• Est. Priority Date: 04/03/2012
• Status: Active Grant

• Current Assignee: Illumina Incorporated
• Original Assignee: Illumina Incorporated
• Inventors: Buermann, Dale, Moon, John A., Crane, Bryan, Wang, Mark, Harris, Jason, Hage, Matthew, Hong, Stanley S., Nibbe, Mark J.
• Primary Examiner(s): CROW, ROBERT THOMAS
• Application Number:
  US14/335,117
• Publication Number:
  US 20140329694A1
• Time in Patent Office: 1,033 Days
• Field of Search: None
• CPC Class Codes: B01L 2200/16 Reagents, handling or storing thereof
• CPC Class Codes: B01L 2300/0877 Flow chambers
• CPC Class Codes: B01L 2400/0487 fluid pressure, pneumatics
• CPC Class Codes: B01L 2400/06 Valves, specific forms thereof
• CPC Class Codes: B01L 3/502715 characterised by interfacing components, e.g. fluidic, electrical, optical or mechanical interfaces
• CPC Class Codes: B01L 3/527 for a plurality of reagents
• 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 2021/0307 Insert part in cell
• CPC Class Codes: G01N 2021/0325 Cells for testing reactions, e.g. containing reagents
• CPC Class Codes: G01N 2021/058 Flat flow cell
• CPC Class Codes: G01N 2021/6417 Spectrofluorimetric devices
• CPC Class Codes: G01N 2021/6441 with two or more labels
• CPC Class Codes: G01N 2021/6482 Sample cells, cuvettes
• CPC Class Codes: G01N 21/6428 Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes" in vivo A61B5/00; immunoassay G01N33/53
• CPC Class Codes: G01N 21/6456 Spatial resolved fluorescence measurements; Imaging
• CPC Class Codes: G01N 21/6486 Measuring fluorescence of biological material, e.g. DNA, RNA, cells G01N21/6428 takes precedence

Abstract

A detection apparatus having a read head including a plurality of microfluorometers positioned to simultaneously acquire a plurality of the wide-field images in a common plane; and (b) a translation stage configured to move the read head along a substrate that is in the common plane. The substrate can be a flow cell that is included in a cartridge, the cartridge also including a housing for (i) a sample reservoir; (ii) a fluidic line between the sample reservoir and the flow cell; (iii) several reagent reservoirs in fluid communication with the flow cell, (iv) at least one valve configured to mediate fluid communication between the reservoirs and the flow cell; and (v) at least one pressure source configured to move liquids from the reservoirs to the flow cell. The detection apparatus and cartridge can be used together or independent of each other.

Claims

1. A method of imaging a substrate, comprising

(a) providing a substrate comprising fluorescent features on a surface;

(b) acquiring a plurality of wide-field images of a first portion of the surface using a co-molded assembly of a plurality of microfluorometers,

wherein each of the microflurometers comprises an objective having a field diameter of at least 1 mm,

wherein each of the microfluorometers acquires a wide-field image from a different location of the surface,

wherein the plurality of microfluorometers are affixed to a carriage; and

(c) translating the carriage in a direction parallel to the surface and repeating (b) for a second portion of the surface.

2. The method of claim 1, further comprising individually focusing each microfluorometer in the plurality of microfluorometers using an autofocus technique.

3. The method of claim 2, wherein the autofocus technique uses at least one dedicated detector that is not used to acquire the wide-field images.

4. The method of claim 2, wherein the autofocus technique uses at least one detector that is also used to acquire the wide-field images.

5. The method of claim 2, wherein the autofocus technique used for a first microfluorometer of the plurality of microfluorometers integrates data from an autofocus technique used for a second microfluorometer of the plurality of microfluorometers, whereby the focus of the first microfluorometer is adjusted based on the focus position of the first microfluorometer and the focus position of the second microfluorometer.

6. The method of claim 1, wherein each of the microfluorometers acquires at least two wide-field images of the surface in (b), each of the at least two wide-field images detecting fluorescence at a different excitation or emission wavelength.

7. The method of claim 1, wherein the fluorescent features have an average center-to-center spacing for nearest neighbors that is less than 50 microns apart and the fluorescent features are individually resolved in the wide-field images.

8. The method of claim 1, wherein each of the microfluorometers acquires a wide-field image of an area of the surface that is at least 1 mm².

9. The method of claim 1, wherein the method further comprises

(d) returning the carriage to a position to acquire a second plurality of wide-field images of the first portion of the surface.

10. The method of claim 9, further comprising modifying the fluorescent features on the surface after (c) and before (d), wherein the second plurality of wide-field images are different from the first plurality of wide-field images.

11. The method of claim 10, wherein the fluorescent features comprise nucleic acids and the modifying comprises altering nucleic acids in a sequencing-by-synthesis technique.

12. The method of claim 10, further comprising repeating (c) a plurality of times prior to (d), thereby imaging at least a third portion and fourth portion of the surface.

13. The method of claim 1, further comprising

(a) providing a fluidic cartridge comprising

(i) a flow cell comprising the surface, wherein the surface is an optically transparent surface,

(ii) a nucleic acid sample,

(iii) a plurality of reagents for a sequencing reaction, and

(iv) a fluidic system for delivering the plurality of reagents to the flow cell;

(b) providing a detection apparatus comprising

(i) the co-molded assembly of the plurality of microfluorometers, and

(ii) a sample stage;

(c) delivering the fluidic cartridge to the sample stage, wherein the optically transparent surface is placed in an image plane having x and y dimensions, and

(d) carrying out fluidic operations of a nucleic acid sequencing procedure in the fluidic cartridge and detection operations of the nucleic acid sequencing procedure in the detection apparatus, wherein

(i) the plurality of reagents are delivered to the flow cell by the fluidic system, and

(ii) the wide-field images of the fluorescent features are detected by the co-molded assembly of the plurality of microfluorometers.