System and method for processing and detecting nucleic acids

• US 9,339,812 B2
• Filed: 02/13/2013
• Issued: 05/17/2016
• Est. Priority Date: 02/13/2012
• Status: Active Grant

• Current Assignee: Neumodx Molecular, Inc. (Qiagen NV)
• Original Assignee: Neumodx Molecular, Inc. (Qiagen NV)
• Inventors: Williams, Jeffrey, Brahmasandra, Sundaresh
• Primary Examiner(s): HORLICK, KENNETH R
• Application Number:
  US13/766,377
• Publication Number:
  US 20130210015A1
• Time in Patent Office: 1,189 Days
• Field of Search: 435/6.12, 435/287.2, 435/288.5
• US Class Current: 1/1
• CPC Class Codes: B01L 2200/025 Align devices or objects to ensure defined positions relative to each other
• CPC Class Codes: B01L 2200/027 for microfluidic devices
• CPC Class Codes: B01L 2200/0605 Metering of fluids
• CPC Class Codes: B01L 2200/0615 Loss of fluid by dripping
• CPC Class Codes: B01L 2200/0668 Trapping microscopic beads
• CPC Class Codes: B01L 2200/0684 Venting, avoiding backpressure, avoid gas bubbles
• CPC Class Codes: B01L 2200/0689 Sealing
• 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 2200/12 Specific details about manufacturing devices
• CPC Class Codes: B01L 2200/142 Preventing evaporation
• CPC Class Codes: B01L 2300/021 Identification, e.g. bar codes
• CPC Class Codes: B01L 2300/022 Transponder chips
• CPC Class Codes: B01L 2300/044 pierceable, e.g. films, membranes
• CPC Class Codes: B01L 2300/06 Auxiliary integrated devices, integrated components
• CPC Class Codes: B01L 2300/0609 Holders integrated in container to position an object
• CPC Class Codes: B01L 2300/0627 Sensor or part of a sensor is integrated
• CPC Class Codes: B01L 2300/0636 Integrated biosensor, microarrays
• CPC Class Codes: B01L 2300/0672 Integrated piercing tool
• CPC Class Codes: B01L 2300/0681 Filter
• CPC Class Codes: B01L 2300/0809 rectangular shaped
• CPC Class Codes: B01L 2300/0816 : Cards, e.g. flat sample carriers usually with flow in two horizontal directions
• 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 2300/087 : Multiple sequential chambers
• CPC Class Codes: B01L 2300/0883 : Serpentine channels
• CPC Class Codes: B01L 2300/0887 : Laminated structure
• CPC Class Codes: B01L 2300/0893 : having a very large number of wells, microfabricated wells
• CPC Class Codes: B01L 2300/123 : Flexible; Elastomeric
• CPC Class Codes: B01L 2300/14 : Means for pressure control
• CPC Class Codes: B01L 2300/16 : Surface properties and coatings
• 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 2300/1822 : using Peltier elements
• CPC Class Codes: B01L 2300/1827 : using resistive heater
• CPC Class Codes: B01L 2400/0406 : capillary forces
• CPC Class Codes: B01L 2400/043 : magnetic forces
• CPC Class Codes: B01L 2400/0478 : pistons
• CPC Class Codes: B01L 2400/0481 : squeezing of channels or chambers
• CPC Class Codes: B01L 2400/0487 : fluid pressure, pneumatics
• CPC Class Codes: B01L 2400/0622 : distribution valves, valves having multiple inlets and/or outlets, e.g. metering valves, multi-way valves
• CPC Class Codes: B01L 2400/065 : sliding valves
• CPC Class Codes: B01L 2400/0655 : pinch valves
• CPC Class Codes: B01L 2400/0694 : vents used to stop and induce flow, backpressure valves
• CPC Class Codes: B01L 2400/086 : using baffles or other fixed flow obstructions
• CPC Class Codes: B01L 3/502 : with fluid transport, e.g. in multi-compartment structures centrifugal-type cuvettes G01N21/07; analysis by separation into components G01N30/00; automatic analysers G01N35/00
• 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 3/502723 : characterised by venting arrangements
• 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 3/502746 : characterised by the means for controlling flow resistance, e.g. flow controllers, baffles B01L3/502738 takes precedence
• CPC Class Codes: B01L 3/502761 : specially adapted for handling suspended solids or molecules independently from the bulk fluid flow, e.g. for trapping or sorting beads, for physically stretching molecules investigating characteristics of particles G01N15/00
• CPC Class Codes: B01L 3/508 : rigid containers not provided for above
• CPC Class Codes: B01L 3/527 : for a plurality of reagents
• CPC Class Codes: B01L 3/567 : Valves, taps or stop-cocks in combination with burettes B01L3/0203; in general F16K
• 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: B29C 65/08 : using ultrasonic vibrations non-plastics element to plastics elements B29C65/645
• CPC Class Codes: B29C 65/484 : Moisture curing adhesives
• CPC Class Codes: B29C 65/606 : the rivets being integral with one of the parts to be joined, i.e. staking
• CPC Class Codes: B29C 66/71 : characterised by the composition of the plastics material of the parts to be joined welding bar compositions B29C65/125
• CPC Class Codes: B29C 66/81423 : being concave
• CPC Class Codes: B29C 66/8322 : Joining or pressing tools reciprocating along one axis
• CPC Class Codes: B29K 2021/00 : Use of unspecified rubbers as moulding material
• CPC Class Codes: B29L 2031/756 : Microarticles, nanoarticles
• CPC Class Codes: C12M 23/42 : Integrated assemblies, e.g. cassettes or cartridges
• CPC Class Codes: C12N 13/00 : Treatment of microorganisms or enzymes with electrical or wave energy, e.g. magnetism, sonic waves
• CPC Class Codes: C12N 15/1013 : by using magnetic beads
• CPC Class Codes: C12Q 1/68 : involving nucleic acids
• CPC Class Codes: C12Q 1/6806 : Preparing nucleic acids for analysis, e.g. for polymerase chain reaction [PCR] assay C12Q1/6804 takes precedence
• CPC Class Codes: C12Q 1/6813 : Hybridisation assays
• CPC Class Codes: C12Q 1/686 : Polymerase chain reaction [PCR]

Abstract

A system and method for processing and detecting nucleic acids from a set of biological samples, comprising: a capture plate and a capture plate module configured to facilitate binding of nucleic acids within the set of biological samples to magnetic beads; a molecular diagnostic module configured to receive nucleic acids bound to magnetic beads, isolate nucleic acids, and analyze nucleic acids, comprising a cartridge receiving module, a heating/cooling subsystem and a magnet configured to facilitate isolation of nucleic acids, a valve actuation subsystem configured to control fluid flow through a microfluidic cartridge for processing nucleic acids, and an optical subsystem for analysis of nucleic acids; a fluid handling system configured to deliver samples and reagents to components of the system to facilitate molecular diagnostic protocols; and an assay strip configured to combine nucleic acid samples with molecular diagnostic reagents for analysis of nucleic acids.

Claims

1. A method for processing and detecting nucleic acids from a biological sample, with a cartridge including a fluidic pathway coupled to a sample port and a reagent port, wherein the method comprises:

aligning the cartridge at a cartridge platform of a molecular diagnostic module, the cartridge platform having a set of slots, and the molecular diagnostic module having a cam module contacting a set of pins aligned with the set of slots;

dispensing the biological sample into the fluidic pathway through the sample port of the cartridge;

moving the cam module by transitioning the actuator into an extended configuration, thereby displacing a subset of the set of pins through the set of slots of the cartridge platform and occluding the fluidic pathway at a subset of occlusion positions to form a first truncated portion of the fluidic pathway for controlling the flow of the biological sample through the fluidic pathway;

separating a volume of nucleic acids from the biological sample within the first truncated portion of the fluidic pathway;

removing a portion of the volume of nucleic acids from the fluidic pathway through at least one of the reagent port and the sample port of the cartridge;

combining the portion of the volume of nucleic acids with a molecular diagnostic reagent to produce a nucleic acid-reagent mixture; and

delivering the nucleic acid-reagent mixture through a second portion of the fluidic pathway to a detection chamber.

2. The method of claim 1, further comprising combining the biological sample with a quantity of magnetic beads, to produce a magnetic bead-sample mixture, prior to dispensing the biological sample into the fluidic pathway through the sample port.

3. The method of claim 2, wherein combining the biological sample with a quantity of magnetic beads further comprises heating the biological sample with the quantity of magnetic beads.

4. The method of claim 2, wherein a supernatant is not removed from the magnetic bead-sample mixture prior to dispensing the biological sample into the fluidic pathway.

5. The method of claim 2, wherein the molecular diagnostic module comprises a magnet, and wherein separating a volume of nucleic acids from the biological sample comprises passing the magnet through one of the set of slots of the cartridge platform, thereby providing a magnetic field at the first truncated portion of the fluidic pathway to capture the magnetic bead-sample within the first truncated portion of the fluidic pathway.

6. The method of claim 5, wherein separating the volume of nucleic acids from the biological sample further comprises dispensing a wash solution through the first truncated portion of the fluidic pathway by a fluid port coupled to the fluidic pathway.

7. The method of claim 6, wherein separating the volume of nucleic acids from the biological sample further comprises dispensing a release solution through the fluidic pathway by the fluid port.

8. The method of claim 7, wherein separating the volume of nucleic acids from the biological sample further comprises heating the first portion of the fluidic pathway to facilitate a pH shift, thus releasing nucleic acids from magnetic beads to produce the volume of nucleic acids.

9. The method of claim 1, wherein combining the portion of the volume of nucleic acids with a molecular diagnostic reagent to produce a nucleic acid-reagent mixture comprises aspirating the nucleic acid-reagent mixture from a well and delivering the nucleic acid-reagent mixture into the well multiple times.

10. The method of claim 1, wherein delivering the nucleic acid-reagent mixture through a second portion of the fluidic pathway comprises delivering the nucleic acid-reagent mixture through the reagent port of the cartridge.

11. The method of claim 1, further comprising generating a set of data based on receiving light from the nucleic acid-reagent mixture through an emission filter.

12. The method of claim 11, wherein generating the set of data further comprises detecting a specific nucleic acid sequence from the nucleic acid-reagent mixture within the detection chamber.

13. The method of claim 1, wherein delivering the nucleic acid-reagent mixture through a second portion of the fluidic pathway to a detection chamber comprises delivering the nucleic acid-reagent mixture through a second portion of a fluidic pathway of a second cartridge.

14. The method of claim 1, wherein at least one of dispensing, separating, removing, combining, and delivering is automatically re-performed for the biological sample in response to an indeterminate result.

15. A method for processing and detecting nucleic acids from a set of biological samples with a cartridge having a set of fluidic pathways defined by an elastomeric layer, the method comprising:

combining each biological sample of the set of biological samples with a quantity of magnetic beads to produce a set of nucleic acid-magnetic bead samples;

aligning the cartridge at a cartridge platform of a molecular diagnostic module, the cartridge platform having a set of slots, and the molecular diagnostic module having a cam module contacting a set of pins aligned with the set of slots;

transferring substantially all of each nucleic acid-magnetic bead sample of the set of nucleic acid-magnetic bead samples to a corresponding fluidic pathway of a set of fluidic pathways;

moving the cam module by transitioning the actuator into an extended configuration, thereby displacing a subset of the set of pins through the set of slots of the cartridge platform, and thereby manipulating the elastomeric layer to occlude at least one fluidic pathway of the set of fluidic pathways at a subset of occlusion positions for controlling a flow through the fluidic pathway; and

detecting nucleic acids using a set of detection chambers coupled to the set of fluidic pathways.

16. The method of claim 15, wherein the method for processing and detecting nucleic acids from a set of biological samples comprises processing and detecting nucleic acids from identical portions of a stock biological sample, such that all biological samples in the set of biological samples are substantially identical in composition.

17. The method of claim 15, wherein combining each biological sample of the set of biological samples with a quantity of magnetic beads to produce a set of nucleic acid-magnetic bead samples further comprises heating the set of nucleic acid-magnetic bead samples.

18. The method of claim 15, wherein combining each biological sample of the set of biological samples with a quantity of magnetic beads comprises combining each biological sample of the set of biological samples with a quantity of magnetic beads treated to be at least one of positively charged, magnetic, paramagnetic, and supraparamgnetic.

19. The method of claim 15, further comprising producing a set of nucleic acid volumes from the set of nucleic acid-magnetic bead samples, prior to detecting nucleic acids using a set of detection chambers.

20. The method of claim 19, wherein producing the set of nucleic acid volumes from the set of nucleic acid-magnetic bead samples comprises:

upon passing a magnet through one of the set of slots of the cartridge platform of the molecular diagnostic module, providing a magnetic field spanning a portion of each fluidic pathway in the set of fluidic pathways, thus capturing the set of nucleic acid-magnetic bead samples;

dispensing a wash solution into each fluidic pathway of the set of fluidic pathways;

dispensing a release solution into each fluidic pathway of the set of fluidic pathways; and

heating each fluidic pathway of the set of fluidic pathways to facilitate a pH shift, thus unbinding nucleic acids from magnetic beads to produce the set of nucleic acid volumes.

21. The method of claim 20, wherein dispensing the wash solution and dispensing a release solution comprises dispensing a wash solution and dispensing a release solution into a fluid port, wherein the fluid port is coupled to each fluidic pathway in the set of fluidic pathways.

22. The method of claim 21, further comprising coupling the fluid port to an external fluid handling system.

23. The method of claim 19, further comprising combining each nucleic acid volume of the set of nucleic acid volumes with a molecular diagnostic reagent of a set of molecular diagnostic reagents prior to detecting nucleic acids using a set of detection chambers.

24. The method of claim 23, wherein all molecular diagnostic reagents of the set of molecular diagnostic reagents are configured to facilitate identical assays.

25. The method of claim 16, further comprising producing a set of nucleic acid volumes from the stock biological sample, and combining the set of nucleic acid volumes with a set of molecular diagnostic reagents, to facilitate running multiple assays from the stock biological sample.

26. The method of claim 19, further comprising aspirating the set of nucleic acid volumes from the set of fluidic pathways, wherein aspirating comprises aspirating the set of nucleic acid volumes from a set of reagent ports, wherein each reagent port of the set of reagent ports is coupled to a corresponding fluidic pathway of the set of fluidic pathways.

27. The method of claim 26, further comprising transferring each of the set of nucleic acid-reagent mixtures into the corresponding fluidic pathway of the set of fluidic pathways, wherein transferring comprises transferring the set of nucleic acid-reagent mixtures back into the set of reagent ports.

28. The method of claim 19, further comprising transmitting light through a set of excitation filters toward the set of nucleic acid-reagent mixtures within the set of detection chambers, and receiving light from the set of nucleic acid-reagent mixtures through a set of emission filters.

29. A method for processing and detecting nucleic acids within a cartridge having a fluidic pathway including a set of occlusion positions defined by an elastomeric layer of the cartridge, the method comprising:

aligning the cartridge at a cartridge platform of a molecular diagnostic module, the cartridge platform having a set of slots, and the molecular diagnostic module having a cam module contacting a set of pins aligned with the set of slots and an actuator that provides relative displacement between the cartridge platform and the set of pins;

moving the cam module by transitioning the actuator into an extended configuration, thereby displacing a first subset of the set of pins through the set of slots of the cartridge platform, and thereby manipulating the elastomeric layer to occlude the fluidic pathway at a first subset of the set of occlusion positions, thus defining a first truncated fluidic pathway passing through a magnetic field for controlling a flow through the fluidic pathway;

capturing a sample of nucleic acids bound to magnetic beads within the first truncated fluidic pathway, by the magnetic field; and

moving the cam module, thereby displacing a second subset of the set of pins through the set of slots of the cartridge platform, and thereby manipulating the elastomeric layer to occlude the fluidic pathway, through the elastomeric layer, at a second subset of the set of occlusion positions, thus defining a second truncated fluidic pathway containing the sample of nucleic acids bound to magnetic beads.

30. The method of claim 29, further comprising:

delivering a wash solution into the second truncated fluidic pathway through a fluid port to facilitate production of a volume of nucleic acids

delivering the volume of nucleic acids through a reagent port coupled to the fluidic pathway;

receiving the volume of nucleic acids combined with a volume of molecular diagnostic reagents to produce a nucleic acid-reagent sample;

occluding the fluidic pathway at a third subset of the set of occlusion positions, thus defining a third truncated fluidic pathway coupled to a detection chamber; and

delivering the nucleic acid-reagent sample, through the third truncated fluidic pathway, to the detection chamber.

31. The method of claim 30, wherein delivering a wash solution comprises delivering a wash solution using a syringe pump.

32. The method of claim 30, wherein at least one of delivering the volume of nucleic acids and delivering the nucleic acid-reagent sample comprises delivering using a multichannel pipette head.

33. The method of claim 30, further comprising:

occluding the fluidic pathway at a fourth subset of the set of occlusion positions, thus defining a fourth truncated fluidic pathway containing the sample of nucleic acids bound to magnetic beads and coupled to the fluid port, and

delivering a release solution into the fourth truncated fluidic pathway through the fluid port to facilitate production of the volume of nucleic acids.

34. The method of claim 30, wherein delivering the nucleic acid-reagent sample to the detection chamber comprises delivering the nucleic acid-reagent sample through the reagent port.