Method of generating droplets

Patent metadata

Field	Value
Publication number	US-10279350-B2
Application number	US-201615365894-A
Country	US
Kind code	B2
Filing date	Nov 30, 2016
Priority date	Sep 23, 2008
Publication date	May 7, 2019
Grant date	May 7, 2019

Abstract

Methods of generating droplets. In an exemplary method, a device including a sample well, a carrier well, a droplet well, and a plurality of microfluidic channels is selected. The microfluidic channels include a first channel, a second channel, and a third channel. A discrete volume of sample-containing fluid is placed into the sample well, and a discrete volume of carrier fluid is placed into the carrier well. A pressure differential is created after placing the discrete volumes, to cause fluid flow. Sample-containing fluid flows from the sample well to a droplet-generation region of the device via the first channel. Carrier fluid flows from the carrier well to the droplet-generation region via the second channel. Sample-containing droplets and carrier fluid flow from the droplet-generation region to the droplet well via the third channel.

First claim

We claim:

1. A method of generating droplets, the method comprising:
selecting a device including a sample well, a carrier well, a droplet well, and a plurality of microfluidic channels, the microfluidic channels including a first channel, a second channel, and a third channel that meet one another at a droplet-generation region, wherein the device includes an upper member and a lower member, wherein the upper member has a base portion and a plurality of protrusions, wherein the protrusions are formed integrally with the base portion and project from a top surface of the base portion, wherein a rim of each well is formed by one of the protrusions, wherein the microfluidic channels are formed in a bottom surface of the base portion, and wherein the lower member is attached to the bottom surface of the base portion and seals a bottom side of each of the microfluidic channels;
placing a discrete volume of sample-containing fluid into the sample well, and a discrete volume of carrier fluid into the carrier well; and
creating a pressure differential after the step of placing, such that (a) sample-containing fluid flows from the sample well to the droplet-generation region of the device via the first channel, (b) carrier fluid flows from the carrier well to the droplet-generation region via the second channel, and (c) sample-containing droplets and carrier fluid flow from the droplet-generation region to the droplet well via the third channel.

2. The method of claim 1 , wherein the step of creating a pressure differential includes a step of applying vacuum to the droplet well.

3. The method of claim 1 , wherein the step of step of creating a pressure differential includes a step of engaging a rim of at least one of the wells with a gasket.

4. The method of claim 3 , wherein the pressure differential is created via a manifold that is engaged with the gasket.

5. The method of claim 1 , wherein the step of creating a pressure differential causes carrier fluid to flow separately to the droplet-generation region via a pair of the microfluidic channels.

6. The method of claim 1 , wherein the device has a plurality of droplet-generation regions each connected to a different sample well, a different carrier well, and a different droplet well, and wherein the step of creating a pressure differential causes droplet generation to occur in parallel in the plurality of droplet-generation regions.

7. The method of claim 1 , wherein the upper member is injection molded as a single piece.

8. A method of generating droplets, the method comprising:
selecting a device including a plurality of emulsion-generation units, each emulsion-generation unit including a sample well, a carrier well, a droplet well, and a plurality of microfluidic channels connecting the wells to a droplet-generation region, wherein the microfluidic channels include at least three channels that meet one another at the droplet-generation region, wherein the device includes an upper member and a lower member, wherein the upper member has a base portion and a plurality of protrusions, wherein the protrusions are formed integrally with the base portion and project from a top surface of the base portion, wherein each well has an upper portion formed by one of the protrusions, wherein the microfluidic channels are formed in a bottom surface of the base portion, and wherein the lower member is attached to the bottom surface of the base portion and seals a bottom side of each of the microfluidic channels;
placing sample-containing fluid into the sample well of each emulsion-generation unit, and carrier fluid into the carrier well of each emulsion-generation unit; and creating a pressure differential in each emulsion-generation unit after the step of placing, such that (a) sample-containing fluid flows from the sample well to the droplet-generation region, (b) carrier fluid flows from the carrier well to the droplet-generation region, and (c) sample-containing droplets and carrier fluid flow from the droplet-generation region to the droplet well;
wherein the step of creating a pressure differential is performed via a manifold, and wherein the manifold is sealed to each droplet well with a gasket that is engaged with a rim of the droplet well.

9. The method of claim 8 , wherein the step of creating a pressure differential includes a step of applying vacuum to each droplet well via the manifold.

10. The method of claim 8 , wherein the step of creating a pressure differential causes all of the emulsion-generation units to generate droplets in parallel.

11. The method of claim 8 , wherein the upper member is injection molded as a single piece.

12. A method of generating droplets, the method comprising:
selecting a device including a body and cover layer, the body being injection molded as a single piece and including a base portion and a plurality of protrusions projecting from a top surface of the base portion, a bottom surface of the base portion having a plurality of microfluidic channels formed therein, the cover layer being attached to the bottom surface of the base portion and sealing a bottom side of each microfluidic channel, the device providing a sample well, a carrier well, and a droplet well, wherein an upper portion of each well is formed by one of the protrusions, and wherein the microfluidic channels include a first channel, a second channel, and a third channel that meet one another at a droplet-generation region;
placing a discrete volume of sample-containing fluid into the sample well, and a discrete volume of carrier fluid into the carrier well; and
creating a pressure differential after the step of placing, such that (a) sample-containing fluid flows from the sample well to the droplet-generation region of the device via the first channel, (b) carrier fluid flows from the carrier well to the droplet-generation region via the second channel, and (c) sample-containing droplets and carrier fluid flow from the droplet-generation region to the droplet well via the third channel.

13. The method of claim 12 , further comprising a step of engaging a rim of the droplet well with a gasket.

14. The method of claim 12 , wherein the step of creating a pressure differential includes a step of applying vacuum to the droplet well via a manifold, and wherein the manifold is sealed to the droplet well using the gasket.

15. The method of claim 12 , wherein the device includes a plurality of droplet wells, and wherein the step of creating a pressure differential includes a step of applying vacuum to the plurality of droplet wells via a manifold.

Assignees

• Bio Rad Laboratories

Inventors

• Hindson Benjamin J
• Ness Kevin D
• Colston Jr Billy W
• Milanovich Fred P
• Masquelier Donald A

Classifications

• B01L2200/0689

  Sealing · CPC title

• B01L2300/0819

  Microarrays; Biochips · CPC title

• B01L3/0241

  Drop counters; Drop formers · CPC title

• B01L2200/0673

  Handling of plugs of fluid surrounded by immiscible fluid · CPC title

• G01N21/49

  within a body or fluid · CPC title