Sample-to-answer microfluidic cartridge

• US 9,895,692 B2
• Filed: 08/05/2015
• Issued: 02/20/2018
• Est. Priority Date: 01/29/2010
• Status: Active Grant

• Current Assignee: Revvity Health Sciences Incorporated (Revvity, Inc.)
• Original Assignee: Micronics Inc. (Sony Group Corp.)
• Inventors: Battrell, C. Frederick, Sprague, Isaac, Bragd, Matthew Scott, Capodanno, Jason
• Primary Examiner(s): Kwak, Dean
• Application Number:
  US14/819,182
• Publication Number:
  US 20160193603A1
• Time in Patent Office: 930 Days
• Field of Search: 422502-505, 436180
• CPC Class Codes: B01L 2200/0684 Venting, avoiding backpressure, avoid gas bubbles
• CPC Class Codes: B01L 2200/14 Process control and prevention of errors
• CPC Class Codes: B01L 2200/16 Reagents, handling or storing thereof
• CPC Class Codes: B01L 2300/044 pierceable, e.g. films, membranes
• CPC Class Codes: B01L 2300/0672 Integrated piercing tool
• 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 2300/087 Multiple sequential chambers
• CPC Class Codes: B01L 2300/0887 Laminated structure
• CPC Class Codes: B01L 2300/123 Flexible; Elastomeric
• CPC Class Codes: B01L 2400/0481 squeezing of channels or chambers
• CPC Class Codes: B01L 2400/0487 fluid pressure, pneumatics
• CPC Class Codes: B01L 2400/0638 membrane valves, flap valves
• 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 3/52 Containers specially adapted for storing or dispensing a reagent B01L3/02 takes precedence; containers for medical or pharmaceutical purposes A61J1/00; containers in general B65D; storing or dispensing test elements G01N33/4875; automated reagent dispensing G01N35/1002
• 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: G01N 21/03 Cuvette constructions
• CPC Class Codes: G01N 21/0332 with temperature control control of temperature G05D23/00; cryostats F17C3/08
• CPC Class Codes: G01N 21/17 Systems in which incident light is modified in accordance with the properties of the material investigated where the material investigated is optically excited causing a change in wavelength of the incident light G01N21/63
• CPC Class Codes: G01N 21/64 : Fluorescence; Phosphorescence
• CPC Class Codes: G01N 21/6428 : Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes" in vivo A61B5/00; immunoassay G01N33/53
• CPC Class Codes: Y10T 137/0318 : Processes

Abstract

A microfluidic cartridge and methods for performing a diagnostic, molecular or biochemical assay thereon, where all dried and/or liquid reagents necessary for the assay are contained in the cartridge and the assay requires only the addition of sample. Pneumohydraulic features, chamber and diaphragm technologies are introduced for overcoming the problems of bubble interference and reagent washout during operation of a microfluidic cartridge. The cartridges are inserted into a host instrument for performance of an assay and the cartridge is supplied as a consumable.

Claims

1. A microfluidic cartridge for assay of a target analyte or analytes in a biological sample, which comprises:

a) a plastic, thermally insulative cartridge body enclosing;

i) a hydraulic works comprising a sample inlet, one or more on-board liquid or dry reagents for an assay, and a wettable downstream microfluidic subcircuit with channels and chambers fluidly connected to said sample inlet and vented at a downstream vent; and

ii) a pneumatic works comprising an inlet or inlets for receiving pneumatic pressure and a pneumatic subcircuit with channels and chambers for conveying said pneumatic pressure therefrom; and

b) an on-board reagent reservoir for dispensing a liquid volume into a microfluidic subcircuit of said hydraulic works;

wherein said reagent reservoir comprises;

i) a duplexedly layered diaphragm sealedly separating a pneumatic chamber of said pneumatic works and a hydraulic chamber of said hydraulic works, said duplexedly layered diaphragm having a first side facing said pneumatic works and a second side facing said hydraulic works, a first layer forming said first side thereof, and a second layer forming said second side thereof, said first and second layers enclosing said liquid volume as a liquid center therebetween;

ii) a fluid outlet for receiving and conveying said liquid volume to said downstream microfluidic subcircuit; and

iii) a sharp disposed in said hydraulic chamber, said sharp for rupturing said second layer and for releasing said liquid volume into said hydraulic works when said duplexedly layered diaphragm is piercingly urged into contact with sharp by application of a pressure differential across said duplexedly layered diaphragm,

wherein said hydraulic works is mounted at a tilt angle theta of 10-35 degrees relative to the ground plane on a tilted stage of a host instrument and at least one hydraulic chamber comprises an outlet and intercommunicating channel positioned superiorly relative to said at least one hydraulic chamber for venting a gas or discharging a bubble from said chamber, and the microfluidic cartridge further comprises;

i) a staging manifold having a plurality of chambers, wherein each said chamber of said plurality of chambers is separated into a hydraulic chamber and a pneumatic chamber by an elastic, energy-storing pneumohydraulic diaphragm sealedly mounted therebetween, such that a liquid volume admitted through an inlet into each said hydraulic chamber in series or in parallel distends each said energy-storing pneumohydraulic diaphragm according to an isobaric pressure proportionate to the displacement volume thereof, and wherein said inlet into each said hydraulic chamber is valvedly closeable for equilibrating said hydraulic pressure throughout said staging manifold after filling is complete; and

ii) a plurality of vented downstream channels in parallel, wherein one said channel of said plurality of channels is in fluidic communication with each said hydraulic chamber of said staging manifold, each said vented downstream channel having a valve for closing during filling and pressurization and for opening during draining and depressurization, whereby said hydraulic pressure of said elastic, pneumohydraulic diaphragm in a distended state is passively converted to the work of advancing a meniscus during initial wetout of said plurality of vented downstream channels in parallel.

2. The microfluidic cartridge of claim 1, wherein said hydraulic works comprises a plurality of wettable downstream microfluidic subcircuits with channels and chambers fluidly connected to said sample inlet and vented at one or more downstream vents, wherein each of said plurality of wettable microfluidic subcircuits is provided with a separate detection chamber.

3. The microfluidic cartridge of claim 1, wherein said duplexedly layered diaphragm is a polyurethane diaphragm, a polyvinylidene chloride diaphragm, a polyolefin diaphragm, a polyester diaphragm, a polyethylene diaphragm, a polyethylene terephthalate diaphragm, a nylon diaphragm, or a laminated or co-extruded combination thereof, and optionally includes a metallized film layer.

4. The microfluidic cartridge of claim 1, wherein each said downstream vented channel comprises an inlet to a microfluidic subcircuit, and wherein each said elastic, energy-storing pneumohydraulic diaphragm is adapted for splitting a liquid volume equally between each said downstream microfluidic subcircuit.

5. The microfluidic cartridge of claim 4, wherein said each microfluidic subcircuit comprises at least one reaction chamber with provision for mixing a liquid reagent, a dry reagent, or a combination thereof, with a liquid sample, and at least one detection chamber interfacing with a detection means for detecting a target analyte or analytes.

6. The microfluidic cartridge of claim 1, wherein said pneumatic chamber of said pneumatic works is vented to atmosphere.

7. The microfluidic cartridge of claim 1, wherein said first layer of said duplexedly layered diaphragm is rupture resistant and said second layer is rupture sensitive.

8. The microfluidic cartridge of claim 7, wherein said first layer is a laminated polymer with outer nylon base layer configured to be puncture resistant and said second layer is a laminated polymer with outer polyethylene terephthalate member, wherein said outer polyethylene terephthalate member is puncture susceptible.

9. The microfluidic cartridge of claim 1, wherein said on-board reagent reservoir releases serial liquid volumes by the action of serial pulses of pneumatic pressure applied thereto.

10. The microfluidic cartridge of claim 1, wherein said liquid volume comprises a liquid reactant, a buffer, a rehydrating fluid, or a diluent, said liquid volume for an assay step selected from rehydrating a dry reagent disposed in a downstream chamber or channel, for rinsing a solid phase, for eluting a target analyte or analytes from a solid phase substrate, for making a dilution, for performing a chromatographic separation, for actuating or stopping a reaction, or for detecting said target analyte or analytes and optionally wherein said liquid volume is degassed and said duplexedly layered diaphragm is gas impervious.

11. The microfluidic cartridge of claim 1, wherein said microfluidic cartridge further comprises:

a first pneumohydraulic diaphragm overlying a first hydraulic chamber and a second pneumohydraulic diaphragm overlying a second hydraulic chamber, said first and second hydraulic chambers having a fluidically interconnecting channel;

a thermal interface for two-zone PCR thermocycling, with first thermal interface of said first hydraulic chamber configured for apposing a first heating element and second thermal interface of said second hydraulic chamber configured for apposing a second heating element;

wherein said first pneumohydraulic diaphragm comprises a pneumatic means for driving reciprocal fluid flow between said first and second hydraulic chambers during PCR amplification, and

wherein said interconnecting channel is configured to be operated at a tilt angle theta of 10-35 degrees so as to reduce interference from bubbles.

12. The microfluidic cartridge of claim 11, wherein said second pneumohydraulic diaphragms is an elastomeric diaphragm and is worked passively by the urging of said first pneumohydraulic diaphragm.

13. The microfluidic cartridge of claim 1, further comprising a detection chamber enclosed on a first opposite side by an optical window and on a second opposite side by a thermo-optical window;

and

wherein said detection chamber is operated at a tilt angle theta of 10-35 degrees so as to flush air and bubbles to a vented port superiorly disposed thereon.