Manipulation of fluids, fluid components and reactions in microfluidic systems

• US 10,675,626 B2
• Filed: 09/17/2019
• Issued: 06/09/2020
• Est. Priority Date: 04/19/2007
• Status: Active Grant

• Current Assignee: Brandeis University, President and Fellows of Harvard College (Harvard Corporation)
• Original Assignee: President and Fellows of Harvard College (Harvard Corporation)
• Inventors: Fraden, Seth, Boukellal, Hakim, Jia, Yanwei, Selimovic, Seila, Rowat, Amy, Agresti, Jeremy, Weitz, David A.
• Primary Examiner(s): Hyun, Paul S
• Application Number:
  US16/573,466
• Publication Number:
  US 20200009570A1
• Time in Patent Office: 266 Days
• Field of Search: None
• CPC Class Codes: B01L 2200/0673 Handling of plugs of fluid surrounded by immiscible fluid
• CPC Class Codes: B01L 2300/0861 Configuration of multiple channels and/or chambers in a single devices
• CPC Class Codes: B01L 2300/087 Multiple sequential chambers
• CPC Class Codes: B01L 2300/0877 Flow chambers
• CPC Class Codes: B01L 2400/0487 fluid pressure, pneumatics
• CPC Class Codes: B01L 2400/0688 surface tension valves, capillary stop, capillary break
• CPC Class Codes: B01L 2400/0694 vents used to stop and induce flow, backpressure valves
• CPC Class Codes: B01L 2400/082 Active control of flow resistance, e.g. flow controllers
• 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 3/502784 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: F17D 1/12 Conveying liquids or viscous products by pressure of another fluid
• CPC Class Codes: G01N 1/28 Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Qmounting specimens on microscopic slides G02B21/34; means for supporting the objects or the materials to be analysed in electron microscopes H01J37/20 ; laboratory gas handling apparatus B01L5/00
• CPC Class Codes: G01N 15/0272 with screening; with classification by filtering B01D takes precedence
• CPC Class Codes: G01N 15/1484 microstructural devices
• CPC Class Codes: G01N 2015/0092 Monitoring flocculation or agglomeration
• CPC Class Codes: Y10T 137/0324 With control of flow by a condition or characteristic of a fluid
• CPC Class Codes: Y10T 137/0391 Affecting flow by the addition of material or energy
• CPC Class Codes: Y10T 137/0396 Involving pressure control
• CPC Class Codes: Y10T 137/2082 Utilizing particular fluid
• CPC Class Codes: Y10T 137/218 Means to regulate or vary operation of device
• CPC Class Codes: Y10T 436/2575 : Volumetric liquid transfer

Abstract

Microfluidic structures and methods for manipulating fluids, fluid components, and reactions are provided. In one aspect, such structures and methods can allow production of droplets of a precise volume, which can be stored/maintained at precise regions of the device. In another aspect, microfluidic structures and methods described herein are designed for containing and positioning components in an arrangement such that the components can be manipulated and then tracked even after manipulation. For example, cells may be constrained in an arrangement in microfluidic structures described herein to facilitate tracking during their growth and/or after they multiply.

Claims

1. A method for partitioning a fluid sample, the method comprising:

providing a microfluidic device having a substrate comprising a plurality of microwells in fluid communication with an inlet;

introducing a first fluid into the inlet of the microfluidic device, the first fluid comprising a biological sample and a plurality of beads, each bead comprising a reactive component for binding a target molecule from the biological sample to the bead;

introducing a second fluid immiscible with the first fluid into the microfluidic device and flowing the second fluid towards each of the plurality of microwells so as to form partitions of fluid in corresponding microwells, wherein at least some of the partitions include

a subvolume of the first fluid,

a portion of the biological sample, and

more than one bead; and

maintaining each partition of fluid in corresponding microwells for detection of contents in each partition of fluid.

2. The method of claim 1, wherein a subset of the partitions do not include any beads and a second subset of the partitions each include a single bead.

3. The method of claim 1, wherein each partition of fluid is separated from the others by the second fluid.

4. The method of claim 1, wherein the second fluid is an oil.

5. The method of claim 1, further comprising monitoring one or more of the partitions of fluid and detecting, with an optical detector, emissions from one or more detectable labels associated with one or more beads.

6. The method of claim 5, further comprising determining the presence of the target molecules based on the detection of emissions.

7. The method of claim 6, wherein the optical detector is a fluorescence detector.

8. The method of claim 7, further comprising measuring, with the fluorescence detector, an intensity of a fluorescent signal emitted from at least one of the detectable labels.

9. The method of claim 7, further comprising capturing one or more images of a fluorescent signal emitted from at least one of the detectable labels.

10. The method of claim 5, wherein, upon a reaction in at least one partition of fluid, the emissions are emitted from one or more of the detectable labels.

11. The method of claim 10, wherein the reaction is a polymerase-chain reaction (PCR).

12. The method of claim 1, wherein the target molecule is a single biological molecule.

13. The method of claim 1, wherein the target molecule is a protein.

14. A method for partitioning a fluid sample, the method comprising:

providing a microfluidic device having a substrate comprising a plurality of regions in fluid communication with an inlet;

introducing an aqueous fluid into the inlet of the microfluidic device, the aqueous fluid comprising a biological sample and a plurality of beads, each bead comprising a reactive component for binding a target protein molecule from the biological sample to the bead;

partitioning the aqueous fluid into a plurality of fluid partitions confined within a plurality of corresponding regions of the substrate by introducing a second fluid immiscible with the aqueous fluid into the microfluidic device and flowing the second fluid towards the plurality of regions, wherein for a subset of the fluid partitions, each fluid partition of the subset comprises a subvolume of the aqueous fluid and more than one bead; and

maintaining each fluid partition of the subset in a corresponding region for detection of contents therein.

15. The method of claim 14, wherein each fluid partition is at least partially surrounded by the second fluid.

16. The method of claim 14, wherein the second fluid comprises an oil.

17. The method of claim 14, further comprising monitoring one or more of the fluid partitions and detecting, with an optical detector, emissions from one or more detectable labels associated with one or more beads and determining the presence of target protein molecules based on the detection of emissions.

18. A method for partitioning a fluid sample, the method comprising:

providing a microfluidic device having a substrate comprising a plurality of microwells in fluid communication with an inlet;

introducing an aqueous fluid into the inlet of the microfluidic device, the aqueous fluid comprising a biological sample and a plurality of beads, each bead comprising a reactive component for binding a target molecule from the biological sample to the bead;

introducing a second fluid immiscible with the aqueous fluid into the microfluidic device and flowing the second fluid towards each of the plurality of microwells so as to form aqueous partitions in corresponding microwells,

wherein at least some of the aqueous partitions include

a subvolume of the first fluid,

at least a portion of the biological sample, and

at least one bead, and

wherein a subset of the aqueous partitions do not include any beads and a second subset of the aqueous partitions include more than one bead; and

maintaining each aqueous partition in corresponding microwells for detection of contents in each aqueous partition.