Analysis devices, kits, and related methods for digital quantification of nucleic acids and other analytes

• US 9,447,461 B2
• Filed: 04/05/2012
• Issued: 09/20/2016
• Est. Priority Date: 03/24/2009
• Status: Active Grant

• Current Assignee: University of Chicago
• Original Assignee: California Institute of Technology, University of Chicago
• Inventors: Ismagilov, Rustem F., Shen, Feng, Kreutz, Jason E., Sun, Bing, Du, Wenbin
• Primary Examiner(s): HURST, JONATHAN M
• Application Number:
  US13/440,371
• Publication Number:
  US 20120264132A1
• Time in Patent Office: 1,629 Days
• US Class Current: 1/1
• CPC Class Codes: B01F 33/304 the mixing being performed in a mixing chamber where the products are brought into contact
• CPC Class Codes: B01L 2200/027 for microfluidic devices
• CPC Class Codes: B01L 2300/0864 comprising only one inlet and multiple receiving wells, e.g. for separation, splitting
• CPC Class Codes: B01L 2300/0896 Nanoscaled
• CPC Class Codes: B01L 3/5025 for parallel transport of multiple samples
• CPC Class Codes: B01L 3/502715 characterised by interfacing components, e.g. fluidic, electrical, optical or mechanical interfaces
• CPC Class Codes: B01L 3/502738 characterised by integrated valves microvalves F16K99/0001
• CPC Class Codes: C12Q 1/6851 Quantitative amplification
• CPC Class Codes: C12Q 2531/119 Strand displacement amplification [SDA]
• CPC Class Codes: C12Q 2537/149 Sequential reactions
• CPC Class Codes: C12Q 2563/159 Microreactors, e.g. emulsion PCR or sequencing, droplet PCR, microcapsules, i.e. non-liquid containers with a range of different permeability's for different reaction components
• CPC Class Codes: C12Q 2565/629 being a microfluidic device

Abstract

Provided are devices and methods for effecting processing of samples, including essentially isothermal amplification of nucleic acids, at multiple reaction locations in a single device. In some embodiments, the disclosed devices and methods provide for effecting parallel sample processing in several hundred locations on a single device.

Claims

1. A method, comprising:

dispersing a first sample that comprises at least one molecule of interest among a plurality of first areas in both a first substrate and a second substrate, at least one of the plurality of first areas containing one and only one molecule of interest, wherein dispersing said first sample comprises exerting the sample through a first conduit in fluidic communication with the plurality of first areas via a first continuous fluidic path within said first and second substrates;

dispersing a reactant material into a plurality of second areas in both said first substrate and said second substrate; and

effecting pairwise placement of at least some of the plurality of first areas into direct fluidic communication with at least some of the plurality of second areas so as to form closed reaction chambers and contact reactant material with the first sample, wherein the pairwise placement comprises relative motion between the first and second substrates which isolates the plurality of first areas disposed in the first substrate from the plurality of first areas disposed in the second substrate.

2. The method of claim 1, wherein (a) the first sample is dispersed among the plurality of first areas at essentially the same time among the plurality of first areas, (b) the reactant material is dispersed among the plurality of second areas at essentially the same time among the plurality of second areas, (c) the pairwise placement of the at least some of the plurality of first areas into direct fluidic communication with at least some of the plurality of second areas occurs at essentially the same time among the majority of any pairs formed by first and second areas.

3. The method of claim 1, wherein the first sample comprises a reagent, and wherein the method further comprises effecting a reaction between the reagent and at least one molecule of interest.

4. The method of claim 3, wherein the reaction comprises nucleic acid amplification.

5. The method of claim 4, wherein the amplification is essentially isothermal.

6. The method of claim 4, further comprising recovering a product of the nucleic acid amplification.

7. The method of claim 3, wherein the pairwise placement is effected by relative motion of the first and second areas.

8. The method of claim 1, further comprising effecting a reaction between the contacted reactant material and the first sample.

9. The method of claim 8, wherein the reactant material comprises an amplification reagent.

10. The method of claim 8, wherein the reaction comprises nucleic acid amplification of the at least one molecule of interest.

11. The method of claim 10, wherein the nucleic acid amplification is essentially isothermal.

12. The method of claim 11, further comprising recovering a product of the nucleic acid amplification.

13. The method of claim 1, wherein the pairwise placement places at least 10 first areas into direct fluidic communication with at least 10 second areas.

14. The method of claim 13, wherein the pairwise placement places at least 50 first areas into direct fluidic communication with at least 50 second areas.

15. The method of claim 1, wherein the pairwise placement is effected by relative motion of the first and second areas.

16. The method of claim 1, wherein dispersing said reactant material comprises exerting the reactant material through a second conduit in fluidic communication with the plurality of second areas via a second continuous fluidic pathway within said first and second substrates.