Method of detecting an analyte

• US 8,088,578 B2
• Filed: 08/24/2009
• Issued: 01/03/2012
• 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: Hua, Zhishan, Pollack, Michael G.
• Primary Examiner(s): FORMAN, BETTY J
• Application Number:
  US12/546,253
• Publication Number:
  US 20090311713A1
• Time in Patent Office: 862 Days
• Field of Search: None
• US Class Current: 435/6.1
• 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 a droplet in a droplet operations gap of a droplet actuator in a detection window comprising an opening or window in a substrate of the droplet actuator, the 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, wherein the magnetic field aggregates the magnetically responsive beads in a region of the droplet which is outside 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, wherein the magnetic field aggregates the magnetically responsive beads in a region of the droplet which is outside the detection window;

(c) detecting a signal from the droplet within the detection window produced by the signal-producing substance to detect the presence, absence and/or quantity of the analyte while the magnetically responsive beads are outside the detection window.

2. The method of claim 1 wherein,

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

and

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.

3. The method of claim 1 wherein the magnetic field is generated by a magnetic field source comprising a fixed permanent magnet.

4. The method of claim 1 wherein the magnetic field is generated by a magnetic field source comprising a moveable permanent magnet.

5. The method of claim 1 wherein the magnetic field is generated by a magnetic field source comprising an electromagnet.

6. The method of claim 1 wherein the magnetic field aggregates the magnetically responsive beads at an edge of the droplet.

7. The method of claim 1 wherein the signal is produced by a fluorescent dye.

8. The method of claim 1 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.

9. The method of claim 1 wherein the detection window is provided in a substrate of a droplet actuator device.

10. The method of claim 9 wherein the droplet actuator comprises a plurality of paths of electrodes associated with the droplet operations substrate, each path associated with

a detection window;

and

a magnetic field in proximity to the path, the magnetic field 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.

11. The method of claim 1 wherein the one or more analytes comprise amplified nucleic acid.

12. The method of claim 1 wherein,

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

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

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.

13. The method of claim 12 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.

14. The method of claim 1 wherein the droplet is retained in place in the detection window by electrowetting while the magnetic field aggregates the magnetically responsive beads in a region of the droplet which is outside the detection window.