Cartridge with lysis chamber and droplet generator

Patent metadata

Field	Value
Publication number	US-9243288-B2
Application number	US-201113250815-A
Country	US
Kind code	B2
Filing date	Sep 30, 2011
Priority date	Sep 23, 2008
Publication date	Jan 26, 2016
Grant date	Jan 26, 2016

Abstract

Systems, including apparatus, methods, compositions, kits, and software, for preparing, reacting, detecting, and/or analyzing samples in droplet-based assay systems, among others. The disclosure emphasizes, but is not limited to, a disposable cartridge with lysis chamber and droplet chamber, particularly for use in droplet-based assays.

First claim

The invention claimed is:

1. A method of nucleic acid amplification, comprising:
purifying a fluid sample in a portion of a disposable, single-use cartridge;
lysing the sample in a portion of the cartridge;
combining the purified, lysed sample with a reagent mixture in a portion of the cartridge;
driving flow of at least one carrier fluid stream and a sample stream to a channel intersection at which droplets containing the sample are generated to form an emulsion, wherein the sample stream includes the purified, lysed sample combined with the reagent mixture; and
cycling the emulsion thermally in a thermocycling instrument to which the emulsion is transferred from the cartridge,
wherein the step of combining includes a step of advancing a first plunger that is operatively connected to a first reservoir containing one of the sample and the reagent mixture, while retracting a second plunger that is operatively connected to a second reservoir containing the other of the sample and the reagent mixture, to move fluid out of the first reservoir and into the second reservoir.

2. The method of claim 1 , wherein the step of purifying includes a step of extracting the sample from a sample collector within the cartridge.

3. The method of claim 1 , wherein the step of purifying includes a step of concentrating the sample within the cartridge.

4. The method of claim 1 , wherein the step of purifying includes a step of separating sample components based on size before the step of lysing.

5. The method of claim 1 , wherein the step of purifying includes a step of separating nucleic acids from other sample components after the step of lysing.

6. The method of claim 1 , wherein the step of lysing includes a step of agitating the sample mechanically.

7. The method of claim 6 , wherein the step of agitating the sample mechanically includes a step of applying magnetic force to a stir element.

8. The method of claim 1 , where the step of cycling includes a step of driving flow of the emulsion along a channel that extends through a plurality of temperature zones.

9. The method of claim 1 , wherein the step of cycling includes a step of holding the emulsion in a container while a temperature of the container is changed over time.

10. The method of claim 1 , wherein the cartridge is a first cartridge, wherein the steps of purifying, lysing, combining, and driving are performed with the first cartridge operatively connected to an instrument that controls the steps of purifying, lysing, combining, and driving, further comprising a step of disconnecting the first cartridge and a step of repeating the steps of purifying, lysing, combining, and driving within a second cartridge operatively connected to the instrument after the first cartridge is disconnected.

11. The method of claim 10 , wherein the instrument supplies fluid to the first and second cartridges during at least one of the steps of purifying, lysing, combining, and driving within each cartridge.

12. The method of claim 11 , wherein the instrument supplies a continuous phase for the emulsion.

13. The method of claim 10 , wherein the instrument performs the step of cycling the emulsion thermally.

14. The method of claim 1 , further comprising a step of providing a cartridge including
a sample chamber configured to receive a sample,
one or more reagent chambers each fluidically connected to the sample chamber and each configured to receive sample-containing fluid from the sample chamber and to combine the sample-containing fluid with a reagent mixture, and
one or more droplet generation regions, each including a channel intersection at which droplets containing the sample are generated to form an emulsion and each fluidically connected to one of the reagent chambers and each configured to receive sample/reagent mixture fluid from one of the reagent chambers and to combine the sample/reagent mixture fluid with a continuous phase to form an emulsion of sample-containing droplets, and
wherein the cartridge is configured to be connected to a reusable instrument that drives operation of the cartridge.

15. The method of claim 14 , wherein the step of providing includes a step of providing a cartridge including a reservoir chamber fluidically connected to the sample chamber and configured to supply a reconstitution fluid to the sample chamber.

16. The method of claim 14 , wherein the step of providing includes a step of providing a cartridge in which the one or more reagent chambers are a plurality of reagent chambers.

17. The method of claim 14 , wherein the step of providing includes a step of providing a cartridge in which the one or more droplet generation regions are a plurality of droplet generation regions.

18. The method of claim 14 , wherein the step of providing includes a step of providing a cartridge in which the sample chamber, the reagent chambers, and the droplet generation regions are fluidically connected to each other by fluid channels defined collectively by a first body portion and a second body portion that are attached to each other, and wherein the fluid channels are created by grooves that are formed in a surface of the first body portion and that are covered by a substantially planar surface of the second body portion.

19. The method of claim 14 , wherein the step of providing includes a step of providing a cartridge including an input chamber configured to receive a continuous phase to be transferred to the droplet generation regions.

20. The method of claim 19 , wherein the step of providing includes a step of providing a cartridge including an outlet chamber configured to receive a portion of the continuous phase that has been transferred out of the input chamber, but that has not been utilized in one of the emulsions.

21. The method of claim 14 , wherein the step of providing includes a step of providing a cartridge including a plurality of droplet chambers each configured to receive one of the generated emulsions.

22. The method of claim 14 , wherein the step of providing includes a step of providing a cartridge including a plurality of plungers configured to be moved to transfer fluid into and out of the chambers.

23. The method of claim 22 , wherein the step of providing includes a step of providing a cartridge in which each plunger is configured to act as a valve by selectively closing an entrance to at least one fluid channel when such plunger is in its most advanced position.

24. The method of claim 14 , wherein the step of providing includes a step of providing a cartridge in which a plurality of connectors carry fluid between at least one chamber of the cartridge and the instrument.

25. The method of claim 14 , wherein the step of providing includes a step of providing a cartridge in which the sample chamber includes an agitation element configured to be agitated by magnetic forces.

26. The method of claim 14 , wherein the step of providing includes a step of providing a cartridge in which the one or more reagent chambers include a plurality of reagent chambers that are fluidically connected to the sample chamber in parallel.

27. The method of claim 14 , wherein the continuous phase includes oil, wherein the step of providing includes a step of providing a cartridge in which the cartridge further comprises at least one oil reservoir fluidically connected to at least one of the reagent chambers and configured to suppl

Assignees

• Ness Kevin Dean
• Burd Samuel
• Hindson Benjamin Joseph
• Belgrader Phillip
• Colston Jr Billy W
• Bio Rad Laboratories

Inventors

• Ness Kevin Dean
• Burd Samuel
• Hindson Benjamin Joseph
• Belgrader Phillip
• Colston Jr Billy W

Classifications

• B01L3/502784 Primary

  specially adapted for droplet or plug flow, e.g. digital microfluidics · CPC title

• B01L2300/041

  Connecting closures to device or container · CPC title

• B29C2045/0079

  applying a coating or covering · CPC title

• B29C45/006

  Joining parts moulded in separate cavities · CPC title

• B01L3/0241

  Drop counters; Drop formers · CPC title