High definition microdroplet printer

• US 10,434,507 B2
• Filed: 10/21/2015
• Issued: 10/08/2019
• Est. Priority Date: 10/22/2014
• Status: Active Grant

• Current Assignee: Regents of the University of California (University of California)
• Original Assignee: Regents of the University of California (University of California)
• Inventors: Abate, Adam R., Sciambi, Adam R., Cole, Russell, Gartner, Zev Jordan
• Primary Examiner(s): Bhat, Narayan K
• Application Number:
  US15/520,056
• Publication Number:
  US 20180056288A1
• Time in Patent Office: 1,448 Days
• CPC Class Codes: B01L 2200/0673 Handling of plugs of fluid surrounded by immiscible fluid
• CPC Class Codes: B01L 2300/0645 Electrodes
• 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 2400/0427 Electrowetting
• CPC Class Codes: B01L 2400/086 using baffles or other fixed flow obstructions
• CPC Class Codes: B01L 3/0268 using pulse dispensing or spraying, eg. inkjet type, piezo actuated ejection of droplets from capillaries
• 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: G01N 15/1433 using image recognition
• CPC Class Codes: G01N 15/1459 the analysis being performed on a sample stream
• CPC Class Codes: G01N 15/1484 microstructural devices
• CPC Class Codes: G01N 15/149 specially adapted for sorting particles, e.g. by their size or optical properties
• CPC Class Codes: G01N 2015/1006 for cytology
• CPC Class Codes: G01N 2035/1034 Transferring microquantities of liquid

Abstract

Methods for delivering discrete entities including, e.g., cells, media or reagents to substrates are provided. In certain aspects, the methods include manipulating and/or analyzing qualities of the entities or biological components thereof. In some embodiments, the methods may be used to create arrays of microenvironments and/or for two and three-dimensional printing of tissues or structures. Systems and devices for practicing the subject methods are also provided.

Claims

1. A method of delivering discrete entities to a substrate, the method comprising:

flowing a plurality of discrete entities through a microfluidic device in a carrier fluid, wherein the discrete entities are insoluble and/or immiscible in the carrier fluid, and wherein the microfluidic device comprises a sorter comprising a flow channel comprising a gapped divider comprising a separating wall which extends less than the complete height of the flow channel;

sorting, via the sorter, one or more of the plurality of discrete entities to be delivered through a delivery orifice to the substrate from the plurality of discrete entities;

directing the carrier fluid and the one or more of the plurality of discrete entities through the delivery orifice to the substrate; and

affixing the one or more of the plurality of discrete entities to the substrate.

2. The method of claim 1, wherein the one or more of the plurality of discrete entities are affixed to the substrate via a force, wherein the force is selected from a gravitational force, an electrical force, a magnetic force, and combinations thereof.

3. The method of claim 2, comprising storing the affixed entity under controlled environmental conditions for a storage period, wherein the force is maintained during the storage period.

4. The method of claim 3, wherein the controlled environmental conditions comprise a constant temperature and/or pressure.

5. The method of claim 2, wherein the force is an electrical force.

6. The method of claim 5, wherein the electrical force is a dielectrophoretic force.

7. The method of claim 1, wherein the discrete entities are droplets.

8. The method of claim 7, wherein the droplets are affixed to the substrate via wetting and/or interfacial tension.

9. The method of claim 7, wherein the droplets comprise an aqueous fluid, which is immiscible with the carrier fluid.

10. The method of claim 9, wherein the substrate comprises on a first surface a layer of fluid which is miscible with the carrier fluid and immiscible with the aqueous fluid, and wherein the droplets are affixed to the first surface of the substrate following introduction into the layer of fluid on the first surface of the substrate.

11. The method of claim 1, wherein the discrete entities have a dimension of from about 1 to 1000 μ

m.

12. The method of claim 11, wherein the discrete entities have a diameter of from about 1 to 1000 μ

m.

13. The method of claim 1, wherein the discrete entities have a volume of from about 1 femtoliter to about 1000 nanoliters.

14. The method of claim 1, wherein the plurality of discrete entities is optically scanned prior to the sorting.

15. The method of claim 14, wherein the sorting is based on results obtained from the optical scan.

16. The method of claim 1, wherein the sorter is an active sorter.

17. The method of claim 16, wherein the sorting comprises sorting via dielectrophoresis.

18. The method of claim 1, wherein the plurality of discrete entities comprises discrete entities which differ in composition.

19. The method of claim 1, wherein the microfluidic device is integrated with an automated system which either:

selectively positions the delivery orifice relative to the substrate, and wherein the method comprises selectively positioning via the automated system the delivery orifice relative to the substrate to selectively deliver the one or more of the plurality of discrete entities to one or more locations on or in proximity to the substrate;

or

selectively positions the substrate relative to the delivery orifice, and wherein the method comprises selectively positioning via the automated system the substrate relative to the delivery orifice to selectively deliver the one or more of the plurality of discrete entities to one or more locations on or in proximity to the substrate.

20. The method of claim 19, wherein the method comprises delivering a first member of the plurality of discrete entities to a first location on or in proximity to the substrate and a second member of the plurality of discrete entities to a second location on or in proximity to the substrate.

21. The method of claim 20, wherein the first and second locations are the same.

22. The method of claim 1, wherein one or more biological assays are performed in one or more of the discrete entities before and/or after delivery to the substrate.

23. The method of claim 1, wherein one or more of the plurality of discrete entities comprises a cell.

24. The method of claim 1, wherein one or more of the plurality of discrete entities comprises a nucleic acid.

25. The method of claim 1, wherein the method comprises encapsulating or incorporating one or more reagents into the plurality of discrete entities.

26. The method of claim 25, wherein the one or more reagents comprise amplification reagents.

27. The method of claim 26, wherein the amplification reagents comprise Polymerase Chain Reaction (PCR) reagents.