Droplet-based particle sorting

• US 9,097,662 B2
• Filed: 10/29/2014
• Issued: 08/04/2015
• Est. Priority Date: 04/18/2006
• Status: Expired due to Fees

• Current Assignee: Advanced Liquid Logic, Inc. (Illumina Incorporated)
• Original Assignee: Advanced Liquid Logic, Inc. (Illumina Incorporated)
• Inventors: Pollack, Michael G., Pamula, Vamsee K., Srinivasan, Vijay
• Primary Examiner(s): Gordon, Brian R
• Application Number:
  US14/526,928
• Publication Number:
  US 20150053562A1
• Time in Patent Office: 279 Days
• Field of Search: 422400-404, 422 63- 681, 422 81- 82, 422/82.01, 422/93, 422502-504, 422551-553, 436/180, 436/43, 436/174, 436/164, 436/158, 436/53, 204/450, 204600-604, 204/607, 204/608
• US Class Current: 1/1
• CPC Class Codes: B01L 2200/0668 Trapping microscopic beads
• CPC Class Codes: B01L 2200/0673 Handling of plugs of fluid surrounded by immiscible fluid
• CPC Class Codes: B01L 2300/0816 Cards, e.g. flat sample carriers usually with flow in two horizontal directions
• 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 2400/0427 Electrowetting
• 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: C12Q 1/6806 Preparing nucleic acids for analysis, e.g. for polymerase chain reaction [PCR] assay C12Q1/6804 takes precedence
• CPC Class Codes: G01N 15/01 specially adapted for biological cells, e.g. blood cells investigating sedimentation of particle suspensions in blood G01N15/05
• CPC Class Codes: G01N 15/1031 by measuring electrical or magnetic effects
• CPC Class Codes: G01N 2015/019 Biological contaminants; Fouling
• CPC Class Codes: G01N 2015/1028 Sorting particles
• CPC Class Codes: G01N 2035/1034 Transferring microquantities of liquid
• CPC Class Codes: G01N 27/447 using electrophoresis
• CPC Class Codes: G01N 27/44717 Arrangements for investigating the separated zones, e.g. localising zones
• CPC Class Codes: G01N 27/44721 by optical means
• CPC Class Codes: G01N 33/543 with an insoluble carrier for immobilising immunochemicals
• CPC Class Codes: G01N 35/10 Devices for transferring samples or any liquids to, in, or from, the analysis apparatus, e.g. suction devices, injection devices G01N35/0099 takes precedence
• CPC Class Codes: Y10T 436/11 : Automated chemical analysis
• CPC Class Codes: Y10T 436/118339 : with formation of a segmented stream
• CPC Class Codes: Y10T 436/25 : including sample preparation
• CPC Class Codes: Y10T 436/2575 : Volumetric liquid transfer

Abstract

The present invention relates to droplet-based particle sorting. According to one embodiment, a droplet microactuator is provided and includes: (a) a suspension of particles; and (b) electrodes arranged for conducting droplet operations using droplets comprising particles. A method of transporting a particle is also provided, wherein the method includes providing a droplet comprising the particle and transporting the droplet on a droplet microactuator.

Claims

1. A method of providing a droplet comprising a single cell, the method comprising:

(a) providing a droplet comprising a suspension of cells on an electrowetting droplet actuator surrounded by an oil medium;

(b) dispensing from the droplet of (a) to provide a dispensed droplet, wherein the dispensing is electrowetting-mediated without the use of dielectrophoresis;

(c) determining whether the dispensed droplet on the electrowetting droplet actuator comprises a single cell; and

(d) sorting the dispensed droplet on the electrowetting droplet actuator based on the results of the determining step using electrowetting-mediated droplet operations.

2. The method of claim 1 further comprising repeating steps 50(b), 50(c), and 50(d).

3. The method of claim 1 wherein the oil medium is substantially immiscible with the droplets of 50(a) and (b).

4. The method of claim 1 wherein step 50(c) is achieved by detecting a number of cells in the dispensed droplet based on a property of and/or signal from the dispensed droplet.

5. The method of claim 4 wherein the detecting step is achieved using a sensor.

6. The method of claim 1, further wherein a single cell determined to be comprised in the dispensed droplet comprises a target cell type, and wherein step 50(c) further comprises detecting the target cell type of the single cell in the dispensed droplet based on a property of and/or signal detected from the dispensed droplet.

7. The method of claim 1 further comprising combining a dispensed droplet determined to comprise a single cell with a separate droplet provided on the droplet actuator to form a combined droplet, wherein the separate droplet comprises a substance which interacts with the single cell.

8. The method of claim 7 wherein the substance which may interact with the single cell is selected from the group consisting of:

beads, cells, microparticles, nanoparticles, drugs, proteins, peptides, antigens, toxins, and antibodies.

9. The method of claim 7 further comprising determining in the combined droplet the presence or quantity of the substance indicative of the single cell'"'"'s response to the substance.

10. The method of claim 9 wherein determining in the combined droplet the presence or quantity of the substance indicative of the single cell'"'"'s response to the substance comprises conducting an assay comprising electrowetting-mediated droplet operations.

11. The method of claim 7 wherein the single cell comprises a T cell.

12. The method of claim 1 further comprising determining if the dispensed droplet comprises multiple cells and combining a dispensed droplet determined to comprise multiple cells with the droplet comprising the suspension of cells.

13. The method of claim 1 further comprising:

(a) determining if the dispensed droplet comprises multiple cells;

(b) splitting a dispensed droplet determined to comprise multiple cells to yield two or more daughter droplets; and

(c) analyzing one or more of the daughter droplets for the presence of a single cell.

14. The method of claim 1 further comprising:

(a) determining if the dispensed droplet comprises multiple cells;

(b) combining a dispensed droplet determined to comprise multiple cells with a buffer droplet to form a combined droplet;

(c) splitting the combined droplet to yield two or more daughter droplets; and

(d) analyzing one or more of the daughter droplets for the presence of a single cell.

15. The method of claim 1 further comprising determining if the dispensed droplet does not contain a single cell and transporting a dispensed droplet determined not to contain a single cell back to the droplet comprising the suspension of cells using electrowetting-mediated droplet operations.

16. The method of claim 1 further comprising determining if the dispensed droplet does not contain a single cell and discarding a dispensed droplet determined not to contain a single cell.

17. The method of claim 1 wherein step 50(c) comprises detecting if a signal indicative of a presence of a single cell is present in the dispensed droplet.

18. The method of claim 17 wherein the signal from a dispensed droplet determined to comprise a single cell comprises a fluorescence from a molecule which binds to the single cell.

19. The method of claim 17 wherein the signal from a dispensed droplet determined to comprise a single cell comprises a radioactive signal.

20. The method of claim 17 wherein the signal from a dispensed droplet determined to comprise a single cell comprises a visual image.

21. The method of claim 1 wherein a dispensed droplet determined to comprise a single cell is transported to an on-droplet microactuator reservoir.

22. The method of claim 1 wherein additional droplet operations are conducted on a dispensed droplet determined to comprise a single cell.

23. The method of claim 1 wherein a dispensed droplet determined to comprise a single cell is further analyzed.