Manipulation of beads in droplets and methods for manipulating droplets

• US 8,809,068 B2
• Filed: 04/15/2010
• Issued: 08/19/2014
• Est. Priority Date: 04/18/2006
• Status: Active Grant

• Current Assignee: Advanced Liquid Logic, Inc. (Illumina Incorporated)
• Original Assignee: Advanced Liquid Logic, Inc. (Illumina Incorporated)
• Inventors: Sista, Ramakrishna, Pamula, Vamsee K., Srinivasan, Vijay, Pollack, Michael G., Eckhardt, Allen
• Primary Examiner(s): Sines, Brian J
• Application Number:
  US12/761,066
• Publication Number:
  US 20100279374A1
• Time in Patent Office: 1,587 Days
• Field of Search: 422/50, 422/68.1, 422/82.01, 422/82.02, 422/100, 436/43, 436/149, 436/150, 436/174, 436/179, 436/180
• US Class Current: 436/180
• CPC Class Codes: B01F 31/65 the materials to be mixed being directly submitted to a pulsating movement, e.g. by means of an oscillating piston or air column
• CPC Class Codes: B01F 33/3021 the components to be mixed being combined in a single independent droplet, e.g. these droplets being divided by a non-miscible fluid or consisting of independent droplets
• CPC Class Codes: B01F 33/3031 using electro-hydrodynamic [EHD] or electro-kinetic [EKI] phenomena to mix or move the fluids
• CPC Class Codes: B01F 33/451 wherein the mixture is directly exposed to an electromagnetic field without use of a stirrer, e.g. for material comprising ferromagnetic particles or for molten metal
• CPC Class Codes: B01L 2200/027 for microfluidic devices
• CPC Class Codes: B01L 2200/0647 Handling flowable solids, e.g. microscopic beads, cells, particles
• CPC Class Codes: B01L 2200/0673 Handling of plugs of fluid surrounded by immiscible fluid
• CPC Class Codes: B01L 2300/06 Auxiliary integrated devices, integrated components
• CPC Class Codes: B01L 2300/0654 Lenses; Optical fibres
• CPC Class Codes: B01L 2300/0816 Cards, e.g. flat sample carriers usually with flow in two horizontal directions
• CPC Class Codes: B01L 2300/0861 Configuration of multiple channels and/or chambers in a single devices
• CPC Class Codes: B01L 2300/0864 comprising only one inlet and multiple receiving wells, e.g. for separation, splitting
• CPC Class Codes: B01L 2300/0867 Multiple inlets and one sample wells, e.g. mixing, dilution
• CPC Class Codes: B01L 2300/089 Virtual walls for guiding liquids
• CPC Class Codes: B01L 2300/1827 using resistive heater
• CPC Class Codes: B01L 2400/0406 capillary forces
• CPC Class Codes: B01L 2400/0415 electrical forces, e.g. electrokinetic
• CPC Class Codes: B01L 2400/0421 electrophoretic flow
• CPC Class Codes: B01L 2400/0424 Dielectrophoretic forces
• CPC Class Codes: B01L 2400/0427 Electrowetting
• CPC Class Codes: B01L 2400/043 : magnetic forces
• CPC Class Codes: B01L 2400/0436 : acoustic forces, e.g. surface acoustic waves [SAW]
• CPC Class Codes: B01L 2400/0442 : thermal energy, e.g. vaporisation, bubble jet
• CPC Class Codes: B01L 3/50273 : characterised by the means or forces applied to move the fluids micropumps F04B19/006, of the membrane type F04B43/043
• 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 7/52 : with provision for submitting samples to a predetermined sequence of different temperatures, e.g. for treating nucleic acid samples amplification or hybridisation processes per se C12Q1/68; controlling sequential reactions for synthesis B01J19/0046
• CPC Class Codes: B01L 7/525 : with physical movement of samples between temperature zones
• CPC Class Codes: G01N 1/38 : Diluting, dispersing or mixing samples
• CPC Class Codes: G01N 2001/386 : Other diluting or mixing processes
• CPC Class Codes: G01N 2035/1046 : Levitated, suspended drops
• CPC Class Codes: G01N 33/54326 : Magnetic particles
• CPC Class Codes: Y10T 436/11 : Automated chemical analysis
• CPC Class Codes: Y10T 436/25 : including sample preparation
• CPC Class Codes: Y10T 436/25625 : Dilution
• CPC Class Codes: Y10T 436/2575 : Volumetric liquid transfer

Abstract

The invention provides a method of dispersing or circulating magnetically responsive beads within a droplet in a droplet actuator. The invention, in one embodiment, makes use of a droplet actuator with a plurality of droplet operations electrodes configured to transport the droplet, and a magnetic field present at a portion of the plurality of droplet operations electrodes. A bead-containing droplet is provided on the droplet actuator in the presence of the uniform magnetic field. Beads are circulated in the droplet during incubation by conducting droplet operations on the droplet within a uniform region of the magnetic field wherein droplet operations do not allow magnetically responsive beads to be introduced into a region of the magnetic field which is sufficiently non-uniform to cause bead clumping resulting in a more homogenous distribution of the beads in the droplet. Other embodiments are also provided.

Claims

1. A method of circulating magnetically responsive beads within a droplet in a droplet actuator, the method comprising:

(a) providing a droplet actuator, comprising;

(i) a plurality of droplet operations electrodes configured to transport the droplet; and

(ii) a magnetic field comprising a uniform region of the magnetic field provided by one or more magnets and present at a plurality of droplet operations electrodes;

(b) providing a bead-containing droplet on the droplet actuator in the presence of the uniform region of the magnetic field; and

(c) circulating beads in the droplet during incubation by conducting droplet transport operations on the droplet using the plurality of droplet operations electrodes within the uniform region of the magnetic field, wherein droplet operations do not allow magnetically responsive beads to be introduced into a region of the magnetic field which is sufficiently non-uniform to cause bead clumping, thereby resulting in a more homogenous distribution of the beads in the droplet, and wherein the droplet is maintained at a temperature suitable for incubation of the contents of the droplet.

2. The method of claim 1 wherein the beads comprise molecules having affinity for one or more target substances in the droplet.

3. The method of claim 1 wherein the beads comprise cells.

4. The method of claim 1 wherein the beads comprise organisms.

5. The method of claim 1 wherein the droplet is surrounded by a filler fluid.

6. The method of claim 1 wherein the method is executed in a manner which avoids clumping of magnetic beads.

7. The method of claim 1 wherein the droplet operations comprise transporting the droplet within the uniform region of the magnetic field.

8. The method of claim 7 wherein the droplet operations comprise transporting the droplet back and forth or in a loop within the uniform region of the magnetic field.

9. The method of claim 7 wherein the droplet operations comprise transporting the droplet back and forth or in a loop within the uniform region of the magnetic field across droplet operations electrodes that vary in size.

10. The method of claim 7 wherein the droplet operations comprise oscillating the droplet within the uniform region of the magnetic field.

11. The method of claim 1 wherein the droplet operations comprise splitting the droplet within the uniform region of the magnetic field.

12. The method of claim 1 wherein the droplet operations comprise splitting and merging the droplet within the uniform region of the magnetic field.

13. The method of claim 1 wherein the droplet operations comprise splitting and merging and transporting the droplet within the uniform region of the magnetic field.

14. The method of claim 1 wherein the droplet is sandwiched between two substrates.

15. The method of claim 1 further comprising splitting the droplet.

16. The method of claim 1 further comprising transporting the droplet away from the magnetic field.

17. The method of claim 1 further comprising splitting the droplet away from the magnetic field.

18. The method of claim 1 further comprising splitting the droplet while proximate an edge of a magnet emanating the magnetic field.

19. The method of claim 1 further comprising recombining the resultant split droplets in the presence of the magnetic field.

20. The method of claim 1 further comprising a split droplet along the plurality of droplet operations electrodes away from the magnetic field.

21. The method of claim 1 further comprising transporting split droplets along the plurality of droplet operations electrodes in opposite directions.

22. The method of claim 1 further comprising producing the magnetic field using at least one magnet of a row of magnets.

23. The method of claim 1 further comprising producing the magnetic field using at least one magnet of a plurality of rows of magnets.