Fluidic devices, systems, and methods for encapsulating and partitioning reagents, and applications of same

• US 10,150,117 B2
• Filed: 09/27/2017
• Issued: 12/11/2018
• Est. Priority Date: 04/10/2014
• Status: Active Grant

• Current Assignee: 10X Genomics Incorporated
• Original Assignee: 10X Genomics Incorporated
• Inventors: Bharadwaj, Rajiv, Ness, Kevin, Mitra, Debkishore, Masquelier, Donald, Makarewicz, Anthony, Hindson, Christopher, Hindson, Benjamin, Saxonov, Serge
• Primary Examiner(s): Bhat, Narayan K
• Application Number:
  US15/717,847
• Publication Number:
  US 20180015473A1
• Time in Patent Office: 440 Days
• 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 2400/049 vacuum
• CPC Class Codes: B01L 2400/086 using baffles or other fixed flow obstructions
• CPC Class Codes: B01L 3/0241 Drop counters; Drop formers making arrays for combinatorial libraries B01J19/0046; automation of dispensing for analysis G01N35/10
• 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: G01N 2035/00237 Handling microquantities of analyte, e.g. microvalves, capillary networks
• CPC Class Codes: G01N 2035/1034 Transferring microquantities of liquid
• CPC Class Codes: G01N 35/085 Flow Injection Analysis

Abstract

The disclosure provides devices, systems and methods for the generation of encapsulated reagents and the partitioning of encapsulated reagents for use in subsequent analyses and/or processing, such as in the field of biological analyses and characterization.

Claims

1. A device for generating a droplet comprising a single microcapsule of a plurality of microcapsules, said device comprising a layered structure comprising:

(i) a first channel connected to a first end of a flow regulator, wherein said first channel is configured to receive a first fluid comprising said plurality of microcapsules, wherein said microcapsules of said first fluid flow at an irregular frequency in said first channel;

(ii) a second channel connected to a droplet generation junction, wherein said second channel is configured to receive a second fluid that is immiscible with said first fluid;

(iii) a third channel connected to a droplet generation junction; and

(iv) a fourth channel connected to a second end of the flow regulator and to the droplet generation junction,

wherein said flow regulator comprises a broadened channel region at said first end and a funneling channel region ending at said second end, wherein said flow regulator is configured to (a) allow microcapsules of said plurality of microcapsules from said first channel to gather in said broadened channel region, and (b) allow said microcapsules gathered in said broadened channel region to flow through said funneling channel region and into said fourth channel at a regular flow frequency,

wherein said fourth channel is configured to bring said microcapsules in contact with said second fluid from said second channel at said droplet generation junction to generate a droplet comprising said single microcapsule from said microcapsules of said plurality of microcapsules, and

wherein said third channel is configured to provide an outlet for said droplet comprising said single microcapsule away from said droplet generation junction.

2. The device of claim 1, wherein said first channel, said second channel, said third channel, and said fourth channel are in a first layer of said layered structure.

3. The device of claim 2, wherein a first reservoir configured to contain said first fluid and a second reservoir configured to contain said second fluid are in a second layer of said layered structure, wherein said second layer is different than said first layer.

4. The device of claim 3, wherein said second layer comprises a planar surface.

5. The device of claim 4, wherein said planar surface of said second layer is mated with a planar surface of said first layer such that said first reservoir is connected to an end of said first channel and said second reservoir is connected to an end of said second channel.

6. The device of claim 5, wherein said first reservoir has a tapering or conical shape towards said end of said first channel.

7. The device of claim 2, wherein said first layer comprises a planar surface in which is disposed a series of channels or grooves that correspond to said first channel, said second channel, said third channel, and said fourth channel.

8. The device of claim 7, further comprising a laminating structure adjacent to said first layer, wherein said laminating structure seals said series of channels or grooves.

9. The device of claim 1, wherein said first channel, said second channel, said third channel, and said fourth channel are injection molded as a single part.

10. The device of claim 1, wherein said flow regulator is dimensioned to provide said microcapsules in said fourth channel at a flow frequency is greater than 50 Hz at a coefficient of variation of less than 30%.

11. The device of claim 1, wherein said flow regulator is dimensioned to provide said microcapsules in said fourth channel at a flow frequency that is sufficient to yield a plurality of droplets at said droplet generation junction, wherein said plurality of droplets has an occupancy of no more than 1 microcapsule per droplet.

12. The device of claim 1, wherein said broadened channel region of said flow regulator narrows along a direction of flow and said funneling channel region rejoins a cross-sectional dimension of said fourth channel.

13. The device of claim 1, wherein said flow regulator comprises a tapering or conical shape.

14. The device of claim 1, wherein said layered structure further comprises a fifth channel, wherein said fifth channel is configured to receive a third fluid comprising an aqueous fluid.

15. The device of claim 14, wherein (i) said first channel or said fourth channel and (ii) said fifth channel meet at a second junction upstream of said droplet generation junction.

16. The device of claim 14, wherein said fifth channel comprises a passive fluid valve that is configured to break capillary forces of an aqueous fluid of said third fluid.

17. The device of claim 1, wherein said first channel or said fourth channel comprises a passive fluid valve that is configured to break capillary forces of an aqueous fluid of said first fluid source.

28. The device of claim 1, wherein said flow regulator is dimensioned to provide said microcapsules in said fourth channel at a flow frequency that is substantially regular in said fourth channel as determined by a number of microcapsules that flow past a given point in said fourth channel within a one second period of time.

18. A device for generating a droplet comprising a single microcapsule of a plurality of microcapsules, comprising

a layered structure comprising:

(i) a first channel configured to receive a first fluid comprising said plurality of microcapsules, wherein said first channel comprises a first region comprising a plurality of pillars that is configured to provide controlled dispensing of said plurality of microcapsules;

(ii) a second channel connected to a droplet generation junction, wherein said second channel is configured to receive a second fluid that is immiscible with said first fluid; and

(iii) a third channel connected to said droplet generation junction;

wherein said first channel is configured to bring microcapsules of said plurality of microcapsules in contact with said second fluid from said second channel at said droplet generation junction to generate a droplet comprising a single microcapsule from said microcapsules of said plurality of microcapsules, and

wherein said third channel is configured to provide an outlet for said droplet comprising said single microcapsule away from said droplet generation junction.

19. The device of claim 18, wherein said first channel further comprises posts or tortuous paths.

20. The device of claim 18, wherein said first channel, said second channel, and said third channel are in a first layer of said layered structure.

21. The device of claim 20, wherein a first reservoir configured to contain said first fluid and a second reservoir configured to contain said second fluid are in a second layer of said layered structure, wherein said second layer is different than said first layer.

22. The device of claim 21, wherein said second layer comprises a planar surface.

23. The device of claim 22, wherein said planar surface of said second layer is mated with a planar surface of said first layer such that said first reservoir is connected to an end of said first channel and said second reservoir is connected to an end of said second channel.

24. The device of claim 20, wherein said first layer comprises a planar surface in which is disposed a series of channels or grooves that correspond to said first channel, said second channel, and said third channel.

25. The device of claim 24, further comprising a laminating structure adjacent to said first layer, wherein said laminating structure seals said series of channels or grooves.

26. The device of claim 18, wherein said layered structure further comprises a fourth channel, wherein said fourth channel is configured to receive a third fluid comprising an aqueous fluid.

27. The device of claim 18, wherein said first channel comprises a passive fluid valve that is configured to break capillary forces of an aqueous fluid of said first fluid.

29. The device of claim 18, wherein said layered structure further comprises a flow regulator, wherein a first end of said flow regulator is connected to said first channel and a second end of said flow regulator is connected to a fourth channel, wherein said flow regulator comprises a broadened channel region at said first end and a funneling channel region ending at said second end, and wherein said fourth channel is connected to said droplet generation junction.

30. The device of claim 29, wherein said flow regulator is configured to (a) allow microcapsules from said first channel to gather in said broadened channel region and (b) allow microcapsules gathered in said broadened channel region to flow through said funneling channel region into said fourth channel at a regular flow frequency.