Manipulation of Beads in Droplets and Methods for Splitting Droplets

• US 20100258441A1
• Filed: 04/15/2010
• Published: 10/14/2010
• Est. Priority Date: 04/18/2006
• Status: Active Grant

• Current Assignee: Duke University
• Original Assignee: Advanced Liquid Logic, Inc. (Illumina Incorporated)
• Inventors: Sista, Ramakrishna, Pamula, Vamsee K., Eckhardt, Allen, Srinivasan, Vijay, Pollack, Michael G.
• Granted Patent: US 8,470,606 B2
• Time in Patent Office: Days
• US Class Current: 204/451
• 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: B01L 2200/027 for microfluidic devices
• 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 3/502746 characterised by the means for controlling flow resistance, e.g. flow controllers, baffles B01L3/502738 takes precedence
• 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 2035/1046 : Levitated, suspended drops
• CPC Class Codes: G01N 35/0098 : involving analyte bound to insoluble magnetic carrier, e.g. using magnetic separation magnetic particles used in immunoassays G01N33/54326; magnetic separation in general B03C
• 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 circulating magnetically responsive beads within a droplet in a droplet actuator. The invention also provides methods for splitting droplets. The invention, in one embodiment, makes use of a droplet actuator with top and bottom substrates, a plurality of magnetic fields respectively present proximate the top and bottom substrates, wherein at least one of the magnet fields is selectively alterable, and a plurality of droplet operations electrodes positioned along at least one of the top and bottom surfaces. A droplet is positioned between the top and bottom surfaces and at least one of the magnetic fields is selectively altered.

Claims

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

(a) providing a droplet actuator, comprising;

(i) top and bottom substrates;

(ii) a plurality of magnetic fields respectively present proximate the top and bottom substrates, wherein at least one of the magnet fields is selectively alterable; and

(iii) a plurality of droplet operations electrodes positioned along at least one of the top and bottom surfaces;

(b) positioning a droplet between the top and bottom surfaces; and

(c) selectively altering at least one of the magnetic fields.

2. The method of claim 1 wherein altering at least one of the magnetic fields further comprises activating an electromagnet proximate at least one of the top and bottom surfaces.

3. The method of claim 1 wherein altering at least one of the magnetic fields further comprises activating and deactivating the magnetic fields in a pattern which causes the beads to circulate within the droplet.

4. The method of claim 1 wherein altering at least one of the magnetic fields further comprises activating an electromagnet proximate only one of the top and bottom surfaces.

5. The method of claim 1 further comprising selectively activating at least one of a plurality of electromagnets positioned proximate the top surface.

6. The method of claim 1 further comprising selectively deactivating at least one of a plurality of electromagnets positioned proximate the top surface.

7. The method of claim 1 further comprising selectively activating at least one of a plurality of electromagnets positioned proximate the bottom surface.

8. The method of claim 1 further comprising selectively deactivating at least one of a plurality of electromagnets positioned proximate the bottom surface.

9. The method of claim 1 wherein altering at least one of the magnetic fields further comprises physically altering a position of a magnet emanating the at least one magnetic field.

10. A method of dispensing a bead-containing sub-droplet, the method comprising:

(a) circulating the beads within the droplet as provided in claim 1; and

(b) using droplet operations to dispense the sub-droplet from the droplet.

11. A method of splitting a droplet comprising a magnetically responsive bead 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 present at the plurality of droplet operations electrodes;

(b) providing a magnetically responsive bead-containing droplet on the droplet actuator;

(c) immobilizing the magnetically responsive bead using the magnetic field; and

(d) using the plurality of droplet operations electrodes to split the droplet into first and second droplets, wherein the magnetically responsive bead remains substantially immobilized.

12. The method of claim 11 further wherein the splitting is facilitated using a hydrophilic patch.

13. The method of claim 11 further comprising using a magnet to generate the magnetic field.

14. The method of claim 11 further comprising using a magnet embedded within a gasket of the droplet actuator to generate the magnetic field.

15. The method of claim 11 further comprising positioning a magnet proximate a gasket of the droplet actuator to generate the magnetic field.

16. The method of claim 11 further comprising using a physical barrier to facilitate splitting of the droplet.

17. The method of claim 11 further comprising using a magnetized physical barrier to facilitate splitting of the droplet.

18. A method of splitting a droplet into two or more sub-droplets, the method comprising:

(a) providing a droplet actuator comprising;

(i) a droplet operations surface comprising droplet operations electrodes, the droplet operations electrodes including one or more intermediate electrodes;

(ii) a top substrate separated from the droplet operations surface to form a gap;

(iii) a physical barrier extending into the gap and constricting the gap in proximity with the one or more intermediate electrodes; and

(iv) a droplet on one or more of the droplet operations electrodes;

(b) activating three or more of the droplet operations including one or more of the intermediate electrodes to elongate the droplet; and

(c) deactivating one or more of the intermediate electrodes to cause the droplet to split into two sub-droplets.

19. The method of claim 18 wherein the barrier is mounted on, and extends into the gap from, the top substrate.

20. The method of claim 18 wherein the barrier produces a magnetic field.

21. The method of claim 18 wherein:

(a) the droplet comprises beads or particles;

(b) the barrier restricts passage of the beads or particles; and

(c) the method yields;

(i) a first sub-droplet comprising substantially all of the beads or particles; and

(ii) a second droplet substantially lacking in beads or particles.