Mesoscale polynucleotide amplification devices

• US 6,660,517 B1
• Filed: 11/08/1996
• Issued: 12/09/2003
• Est. Priority Date: 05/01/1992
• Status: Expired due to Term

• Current Assignee: Trustees Of The University Of Pennsylvania (University of Pennsylvania)
• Original Assignee: Trustees Of The University Of Pennsylvania (University of Pennsylvania)
• Inventors: Kricka, Larry J., Wilding, Peter
• Primary Examiner(s): Marschel, Ardin H.
• Application Number:
  US08/745,771
• Time in Patent Office: 2,587 Days
• Field of Search: 435/6, 435/69.1, 435/91.1, 435/91.2, 435/810, 435/289.1, 435/292.1, 435/303.1, 422/68.1, 422/70.1, 422/129, 436/501, 935/77, 935/78, 935/88
• US Class Current: 435/289.1
• CPC Class Codes: B01D 61/18 Apparatus therefor
• CPC Class Codes: B01D 67/0058 by selective elimination of components, e.g. by leaching
• CPC Class Codes: B01D 67/0072 by deposition from the gaseous phase, e.g. sputtering, CVD, PVD
• CPC Class Codes: B01F 25/41 Mixers of the fractal type
• CPC Class Codes: B01J 19/0046 Sequential or parallel reactions, e.g. for the synthesis of polypeptides or polynucleotides; Apparatus and devices for combinatorial chemistry or for making molecular arrays synthesis methods per se C40B50/00
• CPC Class Codes: B01J 19/0093 Microreactors, e.g. miniaturised or microfabricated reactors laboratory containers with capillary fluid transport in microfabricated channels or chambers B01L3/5027
• CPC Class Codes: B01J 2219/0059 Sequential processes
• CPC Class Codes: B01J 2219/00722 Nucleotides
• CPC Class Codes: B01L 2200/027 for microfluidic devices
• 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 2200/146 Employing pressure sensors
• CPC Class Codes: B01L 2300/0816 Cards, e.g. flat sample carriers usually with flow in two horizontal directions
• 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 2300/1827 using resistive heater
• CPC Class Codes: B01L 2300/1861 using radiation
• CPC Class Codes: B01L 2400/0406 capillary forces
• CPC Class Codes: B01L 2400/0487 fluid pressure, pneumatics
• 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 3/502753 : characterised by bulk separation arrangements on lab-on-a-chip devices, e.g. for filtration or centrifugation separation in general B01D; microapparatus for analysis using electrophoresis G01N27/44791; sample preparation G01N1/28
• 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: B01L 7/525 : with physical movement of samples between temperature zones
• CPC Class Codes: B01L 9/527 : for microfluidic devices, e.g. used for lab-on-a-chip
• CPC Class Codes: C12M 41/46 : of cellular or enzymatic activity or functionality, e.g. cell viability
• CPC Class Codes: C12Q 1/6844 : Nucleic acid amplification reactions
• CPC Class Codes: C12Q 1/686 : Polymerase chain reaction [PCR]
• CPC Class Codes: C40B 40/06 : Libraries containing nucleotides or polynucleotides, or derivatives thereof
• CPC Class Codes: Y10S 366/03 : Micromixers: variable geometry from the pathway influences mixing/agitation of non-laminar fluid flow
• CPC Class Codes: Y10S 435/808 : Optical sensing apparatus
• CPC Class Codes: Y10S 435/81 : Packaged device or kit
• CPC Class Codes: Y10S 435/814 : Enzyme separation or purification
• CPC Class Codes: Y10T 436/25125 : Digestion or removing interfering materials

Abstract

Disclosed are devices for amplifying a preselected polynucleotide in a sample by conducting a polynucleotide amplification reaction. The devices are provided with a substrate microfabricated to include a polynucleotide amplification reaction chamber, having at least one cross-sectional dimension of about 0.1 to 1000 μm. The device also includes at least one port in fluid communication with the reaction chamber, for introducing a sample to the chamber, for venting the chamber when necessary, and, optionally, for removing products or waste material from the device. The reaction chamber may be provided with reagents required for amplification of a preselected polynucleotide. The device also may include means for thermally regulating the contents of the reaction chamber, to amplify a preselected polynucleotide. Preferably, the reaction chamber is fabricated with a high surface to volume ratio, to facilitate thermal regulation.

Claims

1. A device for conducting a polynucleotide amplification reaction, the device comprising:

a sample introduction system for introducing a sample into said device, which comprises an inlet port and a vent;

at least one sample flow channel having a cross-section and extending from said inlet port;

a solid substrate fabricated to include at least one polynucleotide amplification reaction chamber in fluid communication with said flow channel and said vent, said reaction chamber being permanently sealed with a cover and having a cross-section, the cross-section of said reaction chamber having at least one mesoscale dimension of width or depth which is between about 0.1 to 1,000 μ

m, said reaction chamber cross-section and said flow channel cross-section being dissimilar; and

a fluid delivery apparatus for delivering fluid to and receiving fluid from said inlet port, wherein said fluid delivery apparatus interfits with said inlet port, and reversibly seals said inlet port.

2. The device of claim 1 wherein said delivery apparatus comprises a syringe.

3. The device of claim 1 wherein said delivery apparatus comprises a pipette, said pipette comprising a pipette tip provided with an aperture for transferring fluid between said pipette tip and said inlet port.

4. A device for conducting a polynucleotide amplification reaction, the device comprising:

a solid substrate;

a cover disposed over and permanently sealed to said substrate; and

at least one polynucleotide amplification reaction chamber, said reaction chamber having a vent and being fabricated in at least one of said substrate or said cover, said reaction chamber having a cross-section, the cross-section of said reaction chamber having at least one mesoscale dimension of width or depth which is between about 0.1 to 1,000 μ

m.,

wherein said cover comprises;

a cavity for receiving and interfitting with a pipette comprising a pipette tip provided with an aperture; and

a flow channel having a cross-section and communicating between said aperture of said pipette tip and said reaction chamber when said pipette is fitted within said cavity, said reaction chamber cross-section and said flow channel cross-section being dissimilar.

5. The device of claim 4 wherein said cover comprises a transparent material.

6. The device of claim 4 wherein said transparent material comprises a material selected from the group consisting of glasses and an organic polymeric materials.

7. The device of claim 4, wherein said aperture is positioned on a wall of said pipette tip, to permit said pipette tip to move, when said pipette is fitted in said cavity, between:

a first position which permits transfer of fluid from said tip through said aperture and said channel to said reaction chamber; and

a second position which permits said aperture to face a wall of said cavity, thereby to seal said channel and said chamber.