Biosensors for biological or chemical analysis and systems and methods for same

• US 9,990,381 B2
• Filed: 11/25/2014
• Issued: 06/05/2018
• Est. Priority Date: 04/16/2012
• Status: Active Grant

• Current Assignee: Illumina Incorporated
• Original Assignee: Illumina Incorporated
• Inventors: Eltoukhy, Helmy A., Kain, Robert C., Feng, Wenyi, Pratt, Mark, Hirschbein, Bernard, Sabounchi, Poorya
• Primary Examiner(s): Brusca, John S.
• Assistant Examiner(s): Wise, Olivia M
• Application Number:
  US14/552,673
• Publication Number:
  US 20150079596A1
• Time in Patent Office: 1,288 Days
• Field of Search: None
• CPC Class Codes: B01L 2300/0654 Lenses; Optical fibres
• CPC Class Codes: B01L 2300/168 Specific optical properties, e.g. reflective coatings
• CPC Class Codes: B01L 3/502715 characterised by interfacing components, e.g. fluidic, electrical, optical or mechanical interfaces
• CPC Class Codes: C12Q 1/6874 involving nucleic acid arrays, e.g. sequencing by hybridisation
• CPC Class Codes: G01N 15/1404 Handling flow, e.g. hydrodynamic focusing
• CPC Class Codes: G01N 15/1434 Optical arrangements
• CPC Class Codes: G01N 21/6486 Measuring fluorescence of biological material, e.g. DNA, RNA, cells G01N21/6428 takes precedence
• CPC Class Codes: G01N 33/6875 Nucleoproteins
• CPC Class Codes: G06F 16/214 Database migration support

Abstract

Method of analyzing signal data from a biosensor including a detection device having an array of light detectors. The method includes obtaining signal data from the light detectors. The signal data includes light scores that are based on an amount of light detected by the light detectors during a plurality of imaging events. The method also includes analyzing the light scores from a group of light detectors for each of the plurality of the imaging events. The method also includes determining respective crosstalk functions of the light detectors in the group. Each of the crosstalk functions for a corresponding light detector is based on the amount of light detected by other light detectors in the group. The method also includes analyzing the signal data for each of the imaging events using the crosstalk functions to determine characteristics of the analytes-of-interest.

Claims

1. A method of analyzing signal data from a biosensor including a detection device and a flow cell coupled to the detection device, the detection device and the flow cell defining a flow channel therebetween, the detection device including an array of light detectors, each of the light detectors being associated with at least one reaction site having analytes-of-interest, the reaction sites being located along a detector surface of the detection device that defines the flow channel, the method comprising:

illuminating the reaction sites with excitation light, the analytes-of-interest providing light emissions in response to the excitation light, the light emissions propagating through the flow channel and through the detector surface into the detection device;

detecting the light emissions by the light detectors, at least a portion of the light emissions detected by the light detectors including crosstalk light emissions from a non-associated reaction site;

obtaining the signal data from the light detectors, the signal data including light scores that are based on an amount of the light emissions detected by the light detectors during a plurality of imaging events;

analyzing the light scores from a group of light detectors of the array for each of the plurality of the imaging events;

determining respective crosstalk functions of the light detectors in the group from the analyzing the light scores from the group of light detectors of the array for each of the plurality of the imaging events, wherein each of the crosstalk functions for a corresponding light detector in the group is based on the amount of the light emissions detected by other light detectors in the group; and

analyzing the signal data for each of the imaging events using the crosstalk functions to determine characteristics of the analytes-of-interest, wherein;

the detection device includes a filter layer extending between the detector surface and the light detectors, the filter layer including filter walls that are configured to reflect the light emissions toward the light detectors, a light-absorbing material being deposited within chambers that are defined between adjacent filter walls;

the adjacent filter walls define a detection path therebetween through the corresponding light-absorbing material toward the light detectors; and

the filter walls include first and second interior portions, the first interior portion extending in a direction from the reaction sites toward the light detectors, the second interior portion extending in a direction from the first interior portion toward the light detectors.

2. The method of claim 1, wherein the determining respective crosstalk functions of the light detectors in the group includes:

for each imaging event in the plurality of imaging events;

determining a light score for each of the light detectors in the group from the imaging event;

identifying, as emitting, the at least one reaction site associated with the respective light detectors in the group having the light score for the respective imaging event greater than or equal to a predetermined score value;

identifying, as non-emitting, the reaction sites associated with the respective light detectors in the group having the light score for the respective imaging event less than the predetermined score value;

determining that the light score of the light detectors associated with non-emitting reaction sites is entirely from crosstalk emissions; and

determining, for each of the non-emitting reaction sites, the number of adjacent reaction sites identified as emitting and the position of each adjacent reaction site relative to each non-emitting reaction site; and

comparing the light scores of non-emitting reaction sites from the plurality of imaging events to determine crosstalk portions of the light scores provided by the adjacent reaction sites.

3. The method of claim 1, wherein each of the crosstalk functions of the light detectors in the group is based on a number of adjacent reaction sites that provided the light emissions and locations of the adjacent reaction sites that provided the light emissions.

4. The method of claim 1, wherein the analytes-of-interest are nucleic acids.

5. The method of claim 1, wherein the light scores correspond to voltage signals.

6. The method of claim 1, wherein the light emissions detected by the light sensors includes fluorescence emission signals.

7. The method of claim 1, wherein the reaction sites include discrete metal regions on the detector surface having discrete clusters of nucleic acids.

8. The method of claim 1, wherein the imaging events occur according to a predetermined protocol.

9. The method of claim 1, wherein the reaction sites include discrete metal regions on the detector surface.

10. The method of claim 1, wherein the reaction sites include discrete clusters of nucleic acids.

11. The method of claim 1, wherein a density of the reaction sites is at least one million reaction sites per square millimeter.

12. The method of claim 1, wherein the light detectors have detection areas that are less than about 50 μ

m².

13. The method of claim 1, wherein the analytes-of-interest are nucleic acids that form clusters at the reaction sites and the plurality of imaging events occur in accordance with a sequencing-by-synthesis protocol in which fluorescently-labeled nucleotides are incorporated into the nucleic acids prior to each imaging event.

14. The method of claim 1, wherein the detection device includes a passivation layer that defines the detector surface, the analytes-of-interest being immobilized to the detector surface and exposed to the flow channel, the light emissions propagating through the passivation layer toward the light detectors.

15. The method of claim 14, wherein the passivation layer extends between the filter layer and the flow channel, and at least some of the crosstalk light emissions propagate through the flow channel and the passivation layer and over one or more of the filter walls.

16. The method of claim 15, wherein each of the reaction sites corresponding to the group of light detectors is located within a site area along the detector surface, the site areas having shapes defined by the filter walls, wherein the reaction sites have different locations within the site areas.

17. The method of claim 14, wherein:

the passivation layer extends between the filter layer and the flow channel;

the light-absorbing material filters the excitation light; and

the crosstalk light emissions propagate through the flow channel, the passivation layer, and the light-absorbing material to the light detectors.

18. The method of claim 1, wherein

the second interior portion is to attenuate the light emissions from the reaction sites more than the first interior portion.

19. The method of claim 1 wherein:

the first interior portion is coated with a reflective coating material and the second interior portion is not coated or is coated with a different material; and

the light emissions from the reaction sites that are reflected by the coating are less attenuated than the light emissions that are reflected by the second interior portion of the filter walls.

20. The method of claim 1 wherein:

a width for each of the detection paths decreases as the detection path extends toward the light detector;

a thickness of the filter wall begins to increase as the filter wall transitions from the first interior portion to the second interior portion; and

as the thickness of the filter wall increases, the width for the respective detection path decreases.