Method of Detecting an Analyte

• US 20090311713A1
• Filed: 08/24/2009
• Published: 12/17/2009
• Est. Priority Date: 05/13/2008
• Status: Active Grant

• Current Assignee: Advanced Liquid Logic, Inc. (Illumina Incorporated)
• Original Assignee: Advanced Liquid Logic, Inc. (Illumina Incorporated)
• Inventors: Pamula, Vamsee K., Thwar, Prasanna, Eckhardt, Allen E., Srinivasan, Vijay, Pollack, Michael G., Hua, Zhishan
• Granted Patent: US 8,088,578 B2
• Time in Patent Office: Days
• US Class Current: 435/6
• CPC Class Codes: B01L 2200/10 Integrating sample preparation and analysis in single entity, e.g. lab-on-a-chip concept
• CPC Class Codes: B01L 2300/0816 Cards, e.g. flat sample carriers usually with flow in two horizontal directions
• CPC Class Codes: B01L 2300/089 Virtual walls for guiding liquids
• CPC Class Codes: B01L 2400/0427 Electrowetting
• CPC Class Codes: B01L 2400/043 magnetic forces
• CPC Class Codes: B01L 3/502753 characterised by bulk separation arrangements on lab-on-a-chip devices, e.g. for filtration or centrifugation separation in general B01D; microapparatus for analysis using electrophoresis G01N27/44791; sample preparation G01N1/28
• CPC Class Codes: B01L 3/502761 specially adapted for handling suspended solids or molecules independently from the bulk fluid flow, e.g. for trapping or sorting beads, for physically stretching molecules investigating characteristics of particles G01N15/00
• CPC Class Codes: B01L 3/502792 for moving individual droplets on a plate, e.g. by locally altering surface tension
• CPC Class Codes: B01L 7/525 with physical movement of samples between temperature zones
• CPC Class Codes: C12Q 1/6848 characterised by the means for preventing contamination or increasing the specificity or sensitivity of an amplification reaction
• CPC Class Codes: C12Q 2565/629 being a microfluidic device

Abstract

The invention relates to certain novel approaches to reducing or eliminating the movement of contaminants from one droplet to another on a droplet actuator via liquid filler fluid. In one application, a method of detecting an analyte is provided and includes providing in a detection window a droplet including a signal-producing substance indicative of the presence and/or quantity of an analyte and one or more magnetically responsive beads which may interfere with signal produced by the signal producing substance. The method further includes using a magnetic field for magnetically removing the magnetically responsive beads from the detection window and/or magnetically restricting the magnetically responsive beads from entering the detection window while transporting and/or retaining the droplet in the detection window. The method additionally includes detecting a signal produced by the signal-producing substance.

Claims

1. A method of detecting an analyte, the method comprising:

(a) providing in a detection window a droplet comprising;

(i) a signal-producing substance indicative of the presence and/or quantity of an analyte;

(ii) one or more magnetically responsive beads which may interfere with signal produced by the signal producing substance;

(b) using a magnetic field for;

(i) magnetically removing the magnetically responsive beads from the detection window; and

/or

(ii) magnetically restricting the magnetically responsive beads from entering the detection window while transporting and/or retaining the droplet in the detection window;

(c) detecting a signal produced by the signal-producing substance.

2. The method of claim 1 wherein:

(a) the droplet is provided in a droplet operations gap of a droplet actuator; and

(b) the detection window comprises an opening or window in a substrate of the droplet actuator.

3. The method of any of claims 1 and following, wherein:

(a) step 1(b) comprises providing a fixed magnet in proximity to the detection window; and

(b) transporting the droplet into the detection window delivers the magnetically responsive beads into sufficient proximity with the fixed magnet that the beads are pulled away from and/or restricted from entering the detection window.

4. The method of any of the foregoing claims 1 and following, wherein the magnetic field is generated by a magnetic field source comprising a fixed permanent magnet.

5. The method of any of the foregoing claims 1 and following, wherein the magnetic field is generated by a magnetic field source comprising a moveable permanent magnet.

6. The method of any of the foregoing claims 1 and following, wherein the magnetic field is generated by a magnetic field source comprising an electromagnet.

7. The method of any of the foregoing claims 1 and following, wherein the magnetic field aggregates the magnetically responsive beads at an edge of the droplet.

8. The method of any of the foregoing claims 1 and following, wherein the magnetic field aggregates the magnetically responsive beads in a region of the droplet which is outside the detection window.

9. The method of any of the foregoing claims 1 and following, wherein the magnetic field breaks the magnetically responsive beads away from the droplet.

10. The method of any of the foregoing claims 1 and following, wherein the magnetic field breaks the magnetically responsive beads away from the droplet while the droplet is being held in place or moved by electrode mediated forces.

11. The method of any of the foregoing claims 1 and following, wherein the magnetic field attracts the magnetically responsive beads in a manner which pulls them to an edge of the droplet while the droplet is at least partially in the detection window.

12. The method of claim 1 wherein the magnetic field pulls the magnetically responsive beads out of the droplet as the droplet passes over the magnet.

13. The method of any of the foregoing claims 1 and following, wherein the magnetic field pulls the magnetically responsive beads out of the droplet as the droplet approaches a vicinity of the magnet.

14. The method of any of the foregoing claims 1 and following, wherein the magnetic field pulls the magnetically responsive beads out of the droplet as the droplet approaches the detection window.

15. The method of any of the foregoing claims 1 and following, wherein the signal is produced by a fluorescent dye.

16. The method of any of the foregoing claims 1 and following, wherein the magnetic field attracts the magnetically responsive beads in a manner which restricts substantially all of the beads from entering or re-entering the detection window as the droplet is transported into the detection window.

17. The method of any of the foregoing claims 1 and following, wherein the detection window is provided in a substrate of a droplet actuator device.

18. The method of claim 17 wherein the droplet actuator comprises a plurality of paths of electrodes associated with the droplet operations substrate, each path associated with:

(a) a detection window; and

(b) a magnetic field in proximity to the path arranged for;

(i) magnetically removing the magnetically responsive beads from the corresponding detection window; and

/or

(ii) magnetically restricting the magnetically responsive beads from entering the corresponding detection window while transporting into and/or retaining the droplet in the detection window.

19. The method of any of the foregoing claims 1 and following, wherein the droplet emits a signal indicative of the presence, absence and/or quantity of one or more analytes.

20. The method of claim 19 wherein the one or more analytes comprise amplified nucleic acid.

21. The method of any of the foregoing claims 1 and following, wherein:

(a) the detection window is located in a substrate of a droplet actuator;

(b) the droplet actuator comprises temperature control zones along the path of electrodes for conducting a thermal cycling reaction; and

(c) the method comprises;

(i) thermal cycling an amplification-ready droplet to yield an amplified droplet; and

(ii) transporting the amplified droplet into the detection window.

22. The method of claim 21 comprising transporting a droplet comprising magnetically responsive beads along the path of electrodes to the path of electrodes at least partially into the detection window following 1 or more of the thermal cycles.