Thermal reaction device and method for using the same

• US 7,604,965 B2
• Filed: 03/18/2005
• Issued: 10/20/2009
• Est. Priority Date: 04/03/2003
• Status: Active Grant

• Current Assignee: Standard Biotools Incorporated
• Original Assignee: Fluidigm Corporation
• Inventors: Unger, Marc, Facer, Geoffrey, Nassef, Hany Ramez, Lucero, Michael, McBride, Lincoln
• Primary Examiner(s): Kim; Young J
• Application Number:
  US11/084,357
• Publication Number:
  US 20050252773A1
• Time in Patent Office: 1,677 Days
• Field of Search: None
• US Class Current: 435/91.2
• CPC Class Codes: B01L 2200/0642 Filling fluids into wells by specific techniques
• CPC Class Codes: B01L 2200/10 Integrating sample preparation and analysis in single entity, e.g. lab-on-a-chip concept
• CPC Class Codes: B01L 2200/142 Preventing evaporation
• CPC Class Codes: B01L 2200/147 Employing temperature sensors
• CPC Class Codes: B01L 2300/06 Auxiliary integrated devices, integrated components
• CPC Class Codes: B01L 2300/0636 Integrated biosensor, microarrays
• CPC Class Codes: B01L 2300/0816 Cards, e.g. flat sample carriers usually with flow in two horizontal directions
• CPC Class Codes: B01L 2300/0819 Microarrays; Biochips
• CPC Class Codes: B01L 2300/0861 Configuration of multiple channels and/or chambers in a single devices
• CPC Class Codes: B01L 2300/0864 comprising only one inlet and multiple receiving wells, e.g. for separation, splitting
• CPC Class Codes: B01L 2300/0867 Multiple inlets and one sample wells, e.g. mixing, dilution
• CPC Class Codes: B01L 2300/0874 Three dimensional network
• CPC Class Codes: B01L 2300/123 Flexible; Elastomeric
• CPC Class Codes: B01L 2300/1805 Conductive heating, heat from thermostatted solids is conducted to receptacles, e.g. heating plates, blocks
• CPC Class Codes: B01L 2400/0481 squeezing of channels or chambers
• CPC Class Codes: B01L 2400/0487 fluid pressure, pneumatics
• CPC Class Codes: B01L 2400/06 Valves, specific forms thereof
• CPC Class Codes: B01L 2400/0638 membrane valves, flap valves
• CPC Class Codes: B01L 2400/0655 pinch valves
• CPC Class Codes: B01L 3/0248 Prongs, quill pen type dispenser
• CPC Class Codes: B01L 3/5025 : for parallel transport of multiple samples
• CPC Class Codes: B01L 3/5027 : by integrated microfluidic structures, i.e. dimensions of channels and chambers are such that surface tension forces are important, e.g. lab-on-a-chip B01L3/5023 takes precedence; micromixers B01F33/30; microreactors for synthesis B01J19/0093; microcapillary devices in general B81B1/00
• CPC Class Codes: B01L 3/502707 : characterised by the manufacture of the container or its components by shaping or joining plastic parts B29C59/00, B29C65/00; by laminating B32B37/00; manufacture of microstructural devices in general B81C
• CPC Class Codes: B01L 3/502715 : characterised by interfacing components, e.g. fluidic, electrical, optical or mechanical interfaces
• CPC Class Codes: B01L 3/50273 : characterised by the means or forces applied to move the fluids micropumps F04B19/006, of the membrane type F04B43/043
• CPC Class Codes: B01L 3/502738 : characterised by integrated valves microvalves F16K99/0001
• CPC Class Codes: B01L 7/52 : with provision for submitting samples to a predetermined sequence of different temperatures, e.g. for treating nucleic acid samples amplification or hybridisation processes per se C12Q1/68; controlling sequential reactions for synthesis B01J19/0046
• CPC Class Codes: C12Q 1/68 : involving nucleic acids
• CPC Class Codes: C12Q 1/686 : Polymerase chain reaction [PCR]
• CPC Class Codes: G01N 27/453 : Cells therefor

Abstract

An M×N matrix microfluidic device for performing a matrix of reactions, the device having a plurality of reaction cells in communication with one of either a sample inlet or a reagent inlet through a via formed within an elastomeric block of the device. Methods provided include a method for forming vias in parallel in an elastomeric layer of an elastomeric block of a microfluidic device, the method comprising using patterned photoresist masks and etching reagents to etch away regions or portions of an elastomeric layer of the elastomeric block.

Claims

1. A method for performing an analysis of a sample comprising the steps of:

a) providing a microfluidic device comprising;

a sample channel having cross-sectional dimensions between 0.1 and 1000 μ

m,

valves positioned such that the sample channel is partitioned into a plurality of fluidically isolated chambers by activation of said valves, said valves being formed in said microfluidic device by one or more deflectable membrane(s) that can be deflected into said sample channel,

wherein said deflectable membrane is formed from an elastomeric material having a Young'"'"'s modulus in the range 20 Pa to 1 GPa,

wherein said fluidically isolated chambers are not located at junctions formed by the sample channel and other flow channels that are in fluid communication with the sample channel, and

wherein said sample channel has an entrance for introduction of a fluid, but does not have a separate exit

b) introducing a sample fluid comprising reagents for nucleic acid amplification and at least one amplifiable target into said sample channel,

c) simultaneously actuating said valves to produce at least 500 fluidically isolated chambers, wherein at least one amplifiable target is present in at least one chamber,

d) conducting amplification steps simultaneously in said at least 500 fluidically isolated chambers wherein an amplified target is produced in at least one chamber, and

e) detecting said amplified target.

2. The method of claim 1, wherein said amplification steps are polymerase chain reaction (PCR) steps which result in a PCR product being formed.

3. The method of claim 2, wherein said PCR is digital PCR.

4. The method of claim 3, wherein said polymerase chain reaction produces two different color results.

5. The method of claim 1 wherein the sample channel is a branched partitioning channel system and said sample is introduced into said branched partitioning system and a plurality of valves are actuated to isolate said sample into a plurality of isolated reaction volumes.

6. The method of claim 1 wherein the analysis is single nucleotide polymorphism analysis.

7. The method of claim 1 wherein the analysis is genotyping.

8. The method of claim 1 wherein the analysis is real time PCR.

9. The method of claim 1 wherein the fluid capacity of each chamber is less than 20 nL.

10. The method of claim 9 wherein the fluid capacity of each chamber is less than 10 nL.

11. The method of claim 1 wherein sample channel is formed in an elastomeric substrate.

12. The method of claim 1 wherein the and step (c) comprises simultaneously actuating said valves to isolate at least 1000 separate chambers.

13. The method of claim 12 wherein step (c) comprises simultaneously actuating said valves to isolate at least 2000 separate chambers.

14. The method of claim 13 wherein step (c) comprises simultaneously actuating said valves to isolate at least 3000 separate chambers.

15. The method of claim 14 wherein step (c) comprises simultaneously actuating said valves to isolate at least 5000 separate chambers.

16. The method of claim 15 wherein step (c) comprises simultaneously actuating said valves to isolate at least 10,000 separate chambers.

17. The method of claim 1 wherein deflectable membrane is formed from an elastomeric material having a Young'"'"'s modulus in the range 50 Pa to 10 MPa.

18. The method of claim 17 wherein deflectable membrane is formed from an elastomeric material having a Young'"'"'s modulus in the range 100 Pa to 1 MPa.

19. The method of claim 1 wherein the deflectable membrane is formed from polydimethylsiloxane (PDMS).

20. The method of claim 1 wherein the sample fluid comprises at least one blocking reagent selected from the group consisting of a protein and a detergent.

21. The method of claim 1 wherein the amplification steps comprise isothermal amplification.