Multiwell Droplet Actuator, System and Method

• US 20080044893A1
• Filed: 08/16/2007
• Published: 02/21/2008
• 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: Pamula, Vamsee, Srinivasan, Vijay, Pollack, Michael
• Granted Patent: US 7,998,436 B2
• Time in Patent Office: Days
• US Class Current: 435/305.3
• CPC Class Codes: B01J 19/0046 Sequential or parallel reactions, e.g. for the synthesis of polypeptides or polynucleotides; Apparatus and devices for combinatorial chemistry or for making molecular arrays synthesis methods per se C40B50/00
• CPC Class Codes: B01J 2219/00466 in a slurry
• CPC Class Codes: B01J 2219/00495 Means for heating or cooling the reaction vessels
• CPC Class Codes: B01J 2219/00587 High throughput processes
• CPC Class Codes: B01J 2219/00725 Peptides
• CPC Class Codes: B01L 2200/0642 Filling fluids into wells by specific techniques
• CPC Class Codes: B01L 2200/0647 Handling flowable solids, e.g. microscopic beads, cells, particles
• CPC Class Codes: B01L 2300/0819 Microarrays; Biochips
• CPC Class Codes: B01L 2300/0829 Multi-well plates; Microtitration plates
• CPC Class Codes: B01L 2300/0864 comprising only one inlet and multiple receiving wells, e.g. for separation, splitting
• CPC Class Codes: B01L 2300/0887 Laminated structure
• CPC Class Codes: B01L 2300/0893 having a very large number of wells, microfabricated wells
• CPC Class Codes: B01L 2300/18 Means for temperature control
• CPC Class Codes: B01L 2400/0427 Electrowetting
• CPC Class Codes: B01L 2400/043 magnetic forces
• 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 3/502784 specially adapted for droplet or plug flow, e.g. digital microfluidics automatic analysis using a stream of discrete samples in a tube system G01N35/08
• CPC Class Codes: C07K 1/1136 by reversible modification of the secondary, tertiary or quarternary structure, e.g. using denaturating or stabilising agents
• CPC Class Codes: C30B 29/58 Macromolecular compounds
• CPC Class Codes: C30B 7/00 Single-crystal growth from solutions using solvents which are liquid at normal temperature, e.g. aqueous solutions from molten solvents C30B9/00; by normal or gradient freezing C30B11/00; under a protective fluid C30B27/00
• CPC Class Codes: Y10T 436/2575 : Volumetric liquid transfer

Abstract

Multiwell droplet actuators, systems and methods are provided. According to one embodiment, a substrate is provided and comprises: (a) one or more input ports for introduction of one or more reagents and/or samples; (b) a regular array of processing wells; and (c) a network of droplet transport pathways comprising pathways that provide direct or indirect droplet transport from each of the input ports to each of the one or more processing wells. Varying droplet actuators and systems related thereto are also provided.

Claims

1. A substrate comprising:

(a) one or more input ports for introduction of one or more reagents and/or samples;

(b) a regular array of processing wells; and

(c) a network of droplet transport pathways comprising pathways that provide direct or indirect droplet transport from each of the input ports to each of the one or more processing wells.

2. The substrate of claim 1 having footprint dimensions 85.48×

127.76 mm².

3. The substrate of claim 1 comprising 96 wells in an 8 by 12 format having a well-to-well pitch of 9 mm.

4. The substrate of claim 1 comprising 384 wells in a 16 by 24 format having a well-to-well pitch of 4.5 mm.

5. The substrate of claim 1 comprising 1,536 wells in a 32 by 48 format having a well-to-well pitch of 2.25 mm.

6. The substrate of claim 1 comprising 6,144 wells in a 96 by 64 format having a well-to-well pitch of 1.125 mm.

7. The substrate of claim 1 comprising 24,576 wells in a 192 by 128 format having a well-to-well pitch of 0.5625 mm.

8. The substrate of claim 1 comprising 98,304 wells in a 384 by 256 format having a well-to-well pitch of 0.28125 mm.

9. The substrate of claim 1 comprising 393,216 wells in a 768 by 512 format having a well-to-well pitch of 0.140625 mm.

10. The substrate of claim 1 comprising 1,572,864 wells in a 1536 by 1024 format having a well-to-well pitch of 0.0703125 mm.

11. A droplet actuator comprising the substrate of any of claims 1-10 further comprising a top plate mounted parallel to the substrate thereby providing a sealable interior volume between the substrate and the top plate.

12. The droplet actuator of claim 11 wherein the network of droplet transport pathways interconnecting each of the wells is arranged such that a droplet extracted from any well can be transported to any other well via the network.

13. The droplet actuator of claim 11 further comprising one or more ports for inputting a substance into one or more of the wells.

14. The substrate of claim 1 wherein the regular array of microliter-volume or nanoliter-volume wells comprises high volume storage wells and low volume processing wells.

15. The substrate of claim 1 further comprising:

(a) one or more storage wells and one or more supply droplet transport pathways interconnecting the one or more storage wells with the network of droplet transport pathways; and

(b) wherein the regular array of microliter-volume or nanoliter-volume wells comprises processing wells.

16. A droplet actuator comprising the substrate of claim 15 further comprising a top plate mounted parallel to the substrate thereby providing a sealable interior volume between the substrate and the top plate.

17. A system comprising the droplet actuator of claim 16 programmed to automatically extract droplets from the one or more storage wells and transport them to the processing wells to generate an array of screening conditions.

18. The droplet actuator of claim 16 further comprising a means for effecting a localized magnetic field.

19. The droplet actuator of claim 18 wherein the means for effecting the localized magnetic field is arranged to attract magnetic particles in a droplet on the droplet actuator during a splitting operation on the droplet such that upon completion of the splitting operation, at least about 99% of the magnetic particles remain in one droplet.

20. The droplet actuator of claim 19 wherein at least about 99.5% of the magnetic particles remain in one droplet.

21. The droplet actuator of claim 19 wherein at least about 99.9% of the magnetic particles remain in one droplet.

22. A system comprising the droplet actuator of claim 19 and a processor electronically coupled to the droplet actuator, wherein the processor is programmed to instruct the droplet actuator to execute a protocol for washing of the magnetic beads.

23. A system comprising the droplet actuator of claim 19 and a processor electronically coupled to the droplet actuator, wherein the processor is programmed to instruct the droplet actuator to execute an elution protocol.

24. The droplet actuator of claim 16 further comprising a means for localized heating of a processing well.

25. A system comprising the droplet actuator of claim 16 and lacking a means of thermocycling the entire droplet actuator.

26. The droplet actuator of claim 19 further comprising a heating element arranged to provide localized heating of a region of the network of droplet transport pathways.

27. The droplet actuator of claim 26 wherein the heating element is selected from the group consisting of thin film heaters;

resistive trace heaters;

light heaters;

thermal heaters;

joule heating comprising apparatus for passing current through a droplet; and

infrared heaters.

28. A system comprising the droplet actuator of claim 26 and a processor electronically coupled to the droplet actuator and the heating element, wherein the processor is programmed to transport a droplet into proximity with the heating element to effect heating.

29. The system of claim 28 wherein the processor is programmed to repetitively transport a droplet into proximity with one or more of the heating element(s) to subject the droplet to a heating and cooling cycle.

30. The system of claim 28 wherein the processor is programmed to repetitively transport multiple droplets into proximity with one or more of the heating element(s) to subject the droplets to a heating and cooling cycle.

31. The system of claim 29 wherein the droplet comprises a PCR-ready solution, and the repetitive heating results in amplification of a DNA.

32. The system of claim 29 wherein the droplet comprises a PCR-ready solution and the repetitive heating is sufficient to result in amplification of a target DNA if the target DNA is present in the PCR-ready solution.

33. The system of claim 32 wherein the PCR-ready solution is substantially lacking in surfactant.

34. The system of claim 32 wherein the PCR-ready droplet is surrounded by a silicone oil.

35. The droplet actuator of claim 26 comprising two or more of the heating elements spaced to maintain temperature zones on the substrate.

36. The system of claim 29 wherein the processor is programmed to control the temperature of the heating element to effect heating of the droplet but without causing significant vaporization the droplet.