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WO2015017801A1 - Système pour nanoanalyse - Google Patents

Système pour nanoanalyse Download PDF

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Publication number
WO2015017801A1
WO2015017801A1 PCT/US2014/049442 US2014049442W WO2015017801A1 WO 2015017801 A1 WO2015017801 A1 WO 2015017801A1 US 2014049442 W US2014049442 W US 2014049442W WO 2015017801 A1 WO2015017801 A1 WO 2015017801A1
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WO
WIPO (PCT)
Prior art keywords
nanochannels
liquid reservoir
liquid
sample
biopolymers
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Ceased
Application number
PCT/US2014/049442
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English (en)
Other versions
WO2015017801A8 (fr
Inventor
Michael David AUSTIN
Han Cao
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Bionano Genomics Inc
Original Assignee
Bionano Genomics Inc
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Bionano Genomics Inc filed Critical Bionano Genomics Inc
Publication of WO2015017801A1 publication Critical patent/WO2015017801A1/fr
Publication of WO2015017801A8 publication Critical patent/WO2015017801A8/fr
Anticipated expiration legal-status Critical
Ceased legal-status Critical Current

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    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L3/00—Containers or dishes for laboratory use, e.g. laboratory glassware; Droppers
    • B01L3/50—Containers for the purpose of retaining a material to be analysed, e.g. test tubes
    • B01L3/508—Containers for the purpose of retaining a material to be analysed, e.g. test tubes rigid containers not provided for above
    • B01L3/5088—Containers for the purpose of retaining a material to be analysed, e.g. test tubes rigid containers not provided for above confining liquids at a location by surface tension, e.g. virtual wells on plates, wires
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L3/00—Containers or dishes for laboratory use, e.g. laboratory glassware; Droppers
    • B01L3/50—Containers for the purpose of retaining a material to be analysed, e.g. test tubes
    • B01L3/502—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures
    • B01L3/5027—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures 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
    • B01L3/502715—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures 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 characterised by interfacing components, e.g. fluidic, electrical, optical or mechanical interfaces
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L3/00—Containers or dishes for laboratory use, e.g. laboratory glassware; Droppers
    • B01L3/50—Containers for the purpose of retaining a material to be analysed, e.g. test tubes
    • B01L3/502—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures
    • B01L3/5027—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures 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
    • B01L3/50273—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures 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 characterised by the means or forces applied to move the fluids
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L3/00—Containers or dishes for laboratory use, e.g. laboratory glassware; Droppers
    • B01L3/50—Containers for the purpose of retaining a material to be analysed, e.g. test tubes
    • B01L3/502—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures
    • B01L3/5027—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures 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
    • B01L3/502761—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures 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 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
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N15/00—Investigating characteristics of particles; Investigating permeability, pore-volume or surface-area of porous materials
    • G01N15/10—Investigating individual particles
    • G01N15/14—Optical investigation techniques, e.g. flow cytometry
    • G01N15/1468—Optical investigation techniques, e.g. flow cytometry with spatial resolution of the texture or inner structure of the particle
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2200/00—Solutions for specific problems relating to chemical or physical laboratory apparatus
    • B01L2200/02—Adapting objects or devices to another
    • B01L2200/025—Align devices or objects to ensure defined positions relative to each other
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2200/00—Solutions for specific problems relating to chemical or physical laboratory apparatus
    • B01L2200/02—Adapting objects or devices to another
    • B01L2200/026—Fluid interfacing between devices or objects, e.g. connectors, inlet details
    • B01L2200/027—Fluid interfacing between devices or objects, e.g. connectors, inlet details for microfluidic devices
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2200/00—Solutions for specific problems relating to chemical or physical laboratory apparatus
    • B01L2200/04—Exchange or ejection of cartridges, containers or reservoirs
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2200/00—Solutions for specific problems relating to chemical or physical laboratory apparatus
    • B01L2200/06—Fluid handling related problems
    • B01L2200/0647—Handling flowable solids, e.g. microscopic beads, cells, particles
    • B01L2200/0663—Stretching or orienting elongated molecules or particles
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2200/00—Solutions for specific problems relating to chemical or physical laboratory apparatus
    • B01L2200/06—Fluid handling related problems
    • B01L2200/0689—Sealing
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2200/00—Solutions for specific problems relating to chemical or physical laboratory apparatus
    • B01L2200/16—Reagents, handling or storing thereof
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2300/00—Additional constructional details
    • B01L2300/06—Auxiliary integrated devices, integrated components
    • B01L2300/0627—Sensor or part of a sensor is integrated
    • B01L2300/0645—Electrodes
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2300/00—Additional constructional details
    • B01L2300/08—Geometry, shape and general structure
    • B01L2300/0809—Geometry, shape and general structure rectangular shaped
    • B01L2300/0816—Cards, e.g. flat sample carriers usually with flow in two horizontal directions
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2300/00—Additional constructional details
    • B01L2300/08—Geometry, shape and general structure
    • B01L2300/0809—Geometry, shape and general structure rectangular shaped
    • B01L2300/0819—Microarrays; Biochips
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2300/00—Additional constructional details
    • B01L2300/08—Geometry, shape and general structure
    • B01L2300/0896—Nanoscaled
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2400/00—Moving or stopping fluids
    • B01L2400/04—Moving fluids with specific forces or mechanical means
    • B01L2400/0403—Moving fluids with specific forces or mechanical means specific forces
    • B01L2400/0415—Moving fluids with specific forces or mechanical means specific forces electrical forces, e.g. electrokinetic
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2400/00—Moving or stopping fluids
    • B01L2400/04—Moving fluids with specific forces or mechanical means
    • B01L2400/0475—Moving fluids with specific forces or mechanical means specific mechanical means and fluid pressure
    • B01L2400/0487—Moving fluids with specific forces or mechanical means specific mechanical means and fluid pressure fluid pressure, pneumatics
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N15/00—Investigating characteristics of particles; Investigating permeability, pore-volume or surface-area of porous materials
    • G01N2015/0038—Investigating nanoparticles

Definitions

  • the present disclosure relates to the field of nanotechnology and, more specifically, to linearizing molecules in nanofluidic channels.
  • Biopolymers such as genomic DNA, are often in the form of semi-flexible entwined polymeric chains. These macromolecules are normally assumed to have a random coil configuration in free solution. For unmodified dsDNA in biological solution, the persistence length (a parameter defining its rigidity) is typically about 50 nm. In order to achieve consistent and accurate characterization of DNA and other biopolymers, it is desirable that the biopolymer be linearized. Further, to facilitate characterization of macromolecules and biopolymers, such as DNA, sequences or features of the macromolecule may be marked, for example, with fluorescent labeling techniques. However, optical mapping techniques for biopolymers have been hindered by low information density for optical maps, and conventional techniques provide only low-throughput capabilities.
  • a nanofluidic system comprises a cartridge having a first liquid reservoir; a second liquid reservoir; and a plurality of nanochannels establishing a fluid flow pathway between the first and second liquid reservoirs; an electrode associated with the first liquid reservoir and adapted to contact a liquid in said first liquid reservoir; an electrode associated with the second liquid reservoir and adapted to contact a liquid in said second liquid reservoir; wherein the cartridge has a top and a bottom; wherein the first liquid reservoir can be accessed from the top of the cartridge to add liquid into the first liquid reservoir; and wherein the nanochannels include a viewing window visible from the bottom of the cartridge to permit imaging of labeled molecules in the nanochannels.
  • the nanofluidic system comprises at least one fiducial marker detectable from the bottom of the cartridge having a fixed location in relation to the nanochannels.
  • the nanofluidic system further comprises a transition zone interposed in a fluid flow pathway between the first reservoir and the nanochannels, the transition zone comprising structures for at least partially straightening coiled or entangled polymers to facilitate movement of the polymers into the nanochannels in linear form.
  • Some aspects described herein include a device for analyzing biopolymers, comprising a nanofluidic chip having at least 10 parallel nanochannels formed therein; an optically transparent cover sealed to the chip and forming one side of the nanochannels; a carrier into which the chip is mounted, the carrier having an top side and a bottom side; a first liquid reservoir accessible from the top side of the carrier; and a second liquid reservoir; wherein the nanochannels are connected with and provide a fluid pathway between the first and second liquid reservoirs.
  • the device further comprises structure for moving biopolymers from the first liquid reservoir into the nanochannels.
  • the structure for moving biopolymers includes a first electrode in electrical contact with the first liquid reservoir, and a second electrode in electrical contact with the second liquid reservoir, such that upon energization of the first and second electrodes, charged biopolymers in the first liquid reservoir are moved into the nanochannels toward the second liquid reservoir.
  • the charged biopolymers are electrostatically moved into the nanochannels.
  • the structure for moving biopolymers applies hydraulic pressure to the first liquid reservoir.
  • the device further comprises an imaging device adapted to image biopolymers in the nanochannels through the optically transparent cover.
  • the imaging device is adapted to image only a portion of the nanochannels at one time, further comprising scanning structure for changing the portion of the nanochannels being imaged to permit a plurality of images to be obtained that collectively cover a desired imaging region of the nanochannels.
  • the device further comprises one or more controllers in the device that are operatively linked to the structure for moving biopolymers, the scanning structure, and the imaging device, wherein the one or more controllers are programmed to (a) activate the structure for moving biopolymers to move biopolymers into the nanochannels in linearized form; (b) maintain the biopolymers in a fixed location and linearized form in the nanochannels while controlling the scanning structure and imaging device to image the imaging region; and then (c) repeat (a) and (b) one or more times.
  • the device further comprises an indexing structure on the carrier adapted to align the carrier and the chip in a predetermined relationship to the imaging device.
  • the device comprises a plurality of first liquid reservoirs and second liquid reservoirs, wherein the nanochannels are connected with and provide a fluid pathway between the plurality of first liquid reservoirs and the plurality of second liquid reservoirs.
  • the plurality of first liquid reservoirs and the plurality of second liquid reservoirs are arranged in a network.
  • the nanochannels are connected with and provide a fluid pathway between one first liquid reservoir and a plurality of second liquid reservoirs.
  • a plurality of first electrodes are in contact with the first liquid reservoir.
  • the device further comprises a temperature control device in thermal contact with the carrier, the thermal device adapted to maintain the temperature of the carrier at a specified temperature.
  • Another embodiment disclosed herein relates to a method of nanoanalysis comprising providing the nanofiuidic device; adding a sample containing biopolymers to the first liquid reservoir; isolating the first and second liquid reservoirs from the ambient environment; applying a motive force to the first liquid reservoir to move the biopolymers from the first liquid reservoir and into through the at least 10 nanochannels, and into the second liquid reservoir; and capturing an image of at least a portion of the biopolymers in the at least 10 nanochannels through the optically transparent window.
  • the method further comprises adding oil, for example mineral oil, to the sample in the first liquid reservoir and the second liquid reservoir to act as a vapor barrier.
  • the method further comprises controlling the temperature of the carrier to minimize evaporation of the sample.
  • the motive force is generated by a pair of electrodes in contact with the sample in the first liquid reservoir and the second liquid reservoir.
  • the motive force is generated by a pressure differential applied between the first liquid reservoir and the second liquid reservoir.
  • FIG. 1 is a perspective view of an embodiment of a cartridge for nanoanalysis.
  • FIG. 2 is a cutaway view of the cartridge of FIG. 1.
  • FIG. 3A is a perspective view of a portion of an embodiment of a system for nanoanalysis.
  • FIG. 3B is a perspective view of a portion of another embodiment of a system for nanoanalysis.
  • FIG. 4A is a cutaway view of a portion of an embodiment of a system for nanoanalysis.
  • FIG. 4B is a cutaway view of an embodiment of a cartridge for nanoanalysis.
  • FIG. 4C is a cutaway view of another embodiment of a cartridge for nanoanalysis.
  • FIG. 5 is a view of an embodiment of nanostructures for nanoanalysis.
  • FIG. 6 is a perspective view of an embodiment of a system for nanoanalysis.
  • FIG. 7 is a block diagram of an embodiment of a control system for a system for nanoanalysis.
  • FIG. 8 is a flow diagram of a process for imaging linearized molecules as they pass through nanochannels.
  • nanochannel means a region defined by borders. Such borders may be physical, electrical, chemical, magnetic, and the like.
  • nanochannel is used to clarify that certain channels are considered nanoscale in certain dimensions.
  • nanofluidic may mean a fluid system having components whose dimensions are on the nanoscale.
  • nanoanalysis may refer to analysis of a macromolecule or biopolymer, such as DNA or RNA, using a nanoscale structure, such as a nanochannel, e.g., a nanofluidic system.
  • DNA refers to DNA of any length (e.g. , 0.1Kb to 100 megabases).
  • the DNA can be a highly pure preparation, crude, or semi crude material.
  • the DNA can come from any biological source or can be synthetic.
  • sample can include any samples containing biopolymers, for example, bodily fluids such as blood, serum, plasma, sputum, lavage fluid, cerebrospinal fluid, urine, semen, sweat, tears, saliva, and the like, and any biological samples derived therefrom.
  • bodily fluids such as blood, serum, plasma, sputum, lavage fluid, cerebrospinal fluid, urine, semen, sweat, tears, saliva, and the like, and any biological samples derived therefrom.
  • blood blood
  • plasma and “serum”
  • saliva any biological samples derived therefrom.
  • samples expressly encompass fractions or processed portions thereof.
  • sample expressly encompasses a processed fraction or portion derived from the biopsy, swab, smear, etc.
  • FIG. 1 is a perspective view of an embodiment of a cartridge 100.
  • the cartridge 100 has a top 101 , a bottom 102, and an indexing feature 105 formed in or as a part of a perimeter of the cartridge 100.
  • the top 101 includes a concave area 110 in which are located a plurality of sample wells 120.
  • the indexing feature 105 may be defined by a fixed shape or perimeter of the cartridge 100.
  • the cartridge 100 may be adapted for insertion or use in a system or apparatus for performing nanoanalysis in which the orientation of the cartridge is important.
  • the system or apparatus may have a receiving portion for receiving the cartridge 100, where the receiving portion has a size and shape corresponding to the outer perimeter of the cartridge 100, which includes the indexing feature 105, such that the cartridge 100 may only be inserted into the receiving area of the system or apparatus in a particular orientation.
  • the indexing feature 105 may be of any shape, such as a cutout, an angle, a protrusion, or other similar feature so long as the indexing feature results in a perimeter for the cartridge 100 which is asymmetrical about at least 1 axis.
  • the cartridge may have more than one indexing feature 105, which, taken together, form a perimeter of cartridge 100 which is asymmetrical about at least 1 axis.
  • the concave area 1 10 includes a lip 1 1 which extends around the perimeter of the concave area 110 and provides a surface for sealing the concave area, by, for example, receiving a corresponding portion of a nanoanalysis apparatus which has a sealing element. This feature will be described in greater detail below.
  • the plurality of sample wells 120 are defined in part by a well structure 124, and have an opening accessible from the top 101 of the cartridge 100 for adding a sample to each of the plurality of sample wells 120.
  • the well structure 124 forms a portion of the boundary of the sample wells 120, and a portion of the boundary of the sample wells 120 is open in order to receive a sample.
  • the sample wells 120 may be arranged in a grid.
  • the sample wells may be aligned in a number of rows, the rows separated by a portion of well structure 124 in which a nanostructure wall 126 is formed, and the columns being separated by well structure 124 in which the nanochannels 128 are not present.
  • the number of rows and columns may vary. In some embodiments, the number of rows or columns may be columns may be 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 30, 40, 50, or more.
  • one nanostructure wall 126 forms a portion of the boundary of each sample well 120. Specifically, one nanostructure wall 126 is associated with a pair of adjacent sample wells 120 located in adjacent rows, as a the portion of the well structure 124 located therebetween.
  • the well structure 124 and nanostructure walls 126 may advantageously be formed from silicon or a silicon-containing material. Using a silicon chip or a silicon-based material allows for easier, more consistent, and more accurate formation of nanochannels 128 in the nanostructure wall 126.
  • the well structure 124 may be formed of any material in which nanostructures may be formed, such as, for example, germanium, germanium oxide, nitride, molybdenum, molybdenum sulfide, tungsten sulfide, carbon, carbide, glass, quartz, fused silica, or any other suitable material.
  • the well structure 124 and the nanostructure wall 126 may be formed from a single silicon wafer or chip which is attached to a carrier, such as the cartridge 100, for use in an imaging system.
  • a plurality of nanochannels 128 are formed in each nanostructure wall 126.
  • Each nanostructure may include 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, 30, 35, 40, 45, 50, 75, 100, 200, 300, about 500, about 1000, about 2000, about 3000, about 4000, about 5000, about 10000, about 50000, about 100000, or more nanochannels.
  • the nanochannels 128 are formed to create a fluid pathway from one sample well 120 to another sample well 120 in an adjacent row.
  • the plurality of nanochannels 128 can be tunnels, tubes, grooves, or similar structures which form a passage from one sample well 120 to another.
  • the nanochannels 128 may be formed by a variety of processes, such as etching, growing, deposition, or any other suitable method. As will be described herein, the nanochannels 128 may advantageously be formed as a plurality of parallel grooves in the nanostructure wall 126, with the open portion of the groove being directed toward the bottom 102 of the cartridge 100.
  • a chip 103 is connected to the bottom side 102 of the cartridge 100 so as to align with or be underneath the concave area 110.
  • the chip 103 includes the substrate 122, the well structure 124, and the nanostructure wall 126, and other associated components.
  • the well structure 124 may be formed on or attached to a substrate 122.
  • the substrate 122 forms a portion, such as a floor, of the sample well 120.
  • the nanochannels 128 are formed in the nanostructure wall 126 such that one boundary of the nanochannels 128 is the substrate 122.
  • the substrate is advantageously transparent over the channel to allow imaging of the contents of the nanochannels 128 from the bottom 102 of the cartridge 100.
  • the cartridge 100 may have more than one concave area 110, each being aligned or over a chip 103. In this way, the cartridge may have more than one chip 103 for used for nanoanalysis of multiple samples using a single cartridge 100.
  • the well structure 124 may be formed directly on the substrate 122, or the substrate 122 may be adhered to the well structure 124, with the substrate 122 forming a surface, for example, a bottom surface of the sample wells 120.
  • the sample wells 120 are arranged in two rows, with the sample wells 120a being in a first row, and the sample wells 120b being in a second row.
  • a sample well 120a is advantageously located adjacent to a sample well 120b, with the nanostructure wall 126 comprising a plurality of nanochannels 128 dividing the two.
  • sample wells 120b may be shaped differently from sample wells 120a in order visually distinguish sample wells 120a from sample wells 120b.
  • the chip 103 may comprise a network of sample wells 120a and 120b divided by a nanostructure wall 126, arranged in any desirable manner.
  • Sample wells 120a and 120b may be aligned linearly, in a circular pattern, or in any other desirable manner, rather than in a grid of rows and columns as described above.
  • a sample wells 120a may be bounded by more than one nanostructure wall 126 having nanochannels 128 formed therein.
  • one sample well 120 is bounded by more than one sample well 120b, such that a macromolecule or biopolymer placed in the sample well 120a may be migrated into the more than one adjacent sample well 120b.
  • the chip 103 may have a single, circular, square, ovoid, triangular, or other shape sample well 120a bounded by a correspondingly shaped nanostructure wall 126 having nanochannels 128 formed therein, and separating the sample well 120a from surrounding sample wells 120b.
  • a single circular sample well 120a may be surrounded by a circular nanostructure wall 126 such that a macromolecule or biopolymer placed in the sample well 120b may be migrated in a radiating pattern through the nanochannels 128 in the nanostructure wall 126 into a single concentric sample well 120b or into a plurality of surrounding sample wells 120b.
  • the chip 103 may comprise a single sample well 120b separated from one or more sample wells 120b through a nanochannel wall 126 which contains regions of nanochannels 128.
  • the chip 103 comprises a single sample well 120a bounded on one side by a nanochannel wall 126, which has three regions of nanochannels 128 and intervening regions without nanochannels 128.
  • the nanochannel 128 regions are disposed within the nanochannel wall 126 so as to create a flowpath through the nanochannel wall 126 into three corresponding sample wells 120b.
  • this arrangement may be reversed, such that three sample wells 120a are separated from a single sample well 120b by a nanostructure wall 126 having corresponding nanochannel 128 regions.
  • the number and arrangement of sample wells 120a and 120b provided herein is exemplary only, and one of skill in the art would understand that other quantities or arrangements of the sample wells 120a and 120b do not depart from the scope of the present application.
  • the substrate 122 can have fiducial marks (not shown) imprinted, engraved, or otherwise located thereon,
  • the fiducial marks may be located on the underside of the substrate, that is, on the side of the substrate 122 corresponding to the bottom 102 of the cartridge 100, or, in other words, the side of the substrate which is not in contact with the well structure 124.
  • the fiducial marks advantageously provide reference marks for aligning and/or calibrating the illumination and optical imaging devices which can be used to image molecules as they move through the nanochannels 128, as will be described below.
  • FIG. 3A depicts an embodiment of a portion of a system for conducting nanoanalysis.
  • the system has an upper assembly 330 and platform 340.
  • the upper assembly 330 is connected to a nanoanalysis system or apparatus via a hinge or other moveable connection.
  • the hinge connection allows for the upper assembly 330 to be moved away from the platform 340, providing room to insert a cartridge 300 in to a recess 342.
  • the upper assembly 330 may then be moved via the hinge connection to bring the upper assembly 330 into close proximity to the platform 340 and the cartridge 300.
  • the upper assembly 330 is configured to contact the cartridge 300 at least at a lip 315, and can form a seal with the lip 315.
  • the upper assembly 330 provides a sealing surface (not shown) which interacts with the lip 315 to create an air and/or water tight seal around the concave portion 310.
  • the platform 340 is formed with a recess 342 configured to receive the cartridge 300.
  • the recess 342 is sized and shaped to correspond to the size and shape of the cartridge 300, such that the cartridge 300 maybe inserted into the recess 342.
  • the recess has an indexing feature 345 which corresponds to an indexing feature 305 on the cartridge 300 to ensure proper orientation and/or alignment within the recess 342 and platform 340.
  • the upper assembly 330 includes a motive force generator.
  • the motive force generator comprises a pressure generation element (not shown), which is configured to apply a pressure gradient or difference between adjacent sample wells.
  • the upper assembly 330 may provide a sealing element (not shown), in order to pressurize the individual sample wells 120a.
  • the pressure gradient can be sufficient to drive molecules from one sample well 120a to the adjacent sample well 120b through the nanostructure wall 126.
  • the pressure generating element may provide a pneumatic pressure, a hydraulic pressure, or other suitable pressure to the sample wells 120a.
  • FIG. 3B depicts another embodiment of a portion of a system for conducting nanoanalysis having a temperature control device 350.
  • the temperature control device 350 comprises a reservoir 352 in contact with the platform 340.
  • the platform 340 can be constructed of a thermally conductive material, such as aluminum.
  • a seal (not shown) between the platform 340 and the temperature control device prevents leakage of a cooling medium out of the reservoir.
  • the temperature control device 350 comprises a cooling inlet 351 which provides a flow of cooling medium into the reservoir 352.
  • the reservoir 352 is in fluid communication with a cooling outlet 353.
  • temperature control device 350 is described here as having cooling inlet 351 and cooling outlet 353, it will be understood by one of skill in the art that temperature control device 350 can circulate a warming medium in order to raise temperature of the platform 340, and not only a cooling medium.
  • cooling medium is used to describe fluids which operate either to add heat to or remove heat from the platform 340.
  • a temperature controller controls a pump or other motive force (not shown) which moves a cooling medium into cooling inlet 351, into the reservoir, where the cooling medium either absorbs heat from, or transfers heat to, the platform 340.
  • the cooling medium exits the reservoir 352 via cooling outlet 353.
  • the cooling medium may be water, air, glycol-based coolant, inert gas, or any other coolant known in the art.
  • a temperature controller can be programmed to maintain the platform at a specified temperature. As the cartridge 300 is in contact with the platform 340, the temperature of the platform helps to control the temperature of the cartridge 300 at a desired level.
  • the temperature controller receives a temperature signal from the reservoir 352 or from the platform 340, or both, as a feedback temperature for controlling the temperature of the sample in cartridge 300.
  • the temperature controller can be programmed to maintain the temperature of the cartridge 300 and/or platform 340 at or near the dew point of the sample, based on atmospheric humidity, temperature, and pressure. In this way, evaporation or condensation of the sample can be reduced, minimized or prevented.
  • the temperature controller may control the temperature of the sample at any desired temperature according to the requirements of the nanoanalysis being performed.
  • FIG. 4A depicts a cutaway view of an embodiment of a portion of a system and a cartridge used for nanoanalysis.
  • a cartridge 400 is depicted in contact with an upper assembly 430.
  • the upper assembly 430 is in contact with a concave portion 410.
  • the cartridge 400 is attached to a chip 403.
  • the chip 403, sample wells 420, well structure 424, and substrate 422 may be similar to those described elsewhere herein.
  • the motive force may be an electric field generated by one or more electrodes.
  • the upper assembly 430 includes negative electrodes 435 and positive electrodes 436 which are supported by the upper assembly 430 and extend downward from the upper assembly 430.
  • the negative electrodes 435 and positive electrodes 436 are connected to the upper assembly 430 such that an end of each electrode 435 and 436 is positioned to align with one sample well 420a or 420b.
  • one negative electrode 435 will be positioned within one sample well 420a and one positive electrode 436 will be positioned within one sample well 420b.
  • This arrangement allows for the creation of an electric field across the nanostructure wall 426, and the electric field can electrophoretically drive biopolymers or macromolecules, such as DNA, from the sample well 420a, through the nanochannels 428 formed in the nanostructure wall 426, and into the adjacent sample well 420b.
  • DNA is generally negatively charged
  • DNA can be moved from one sample well 420a to another sample well 420b through nanochannels 428 toward the positive electrode 436.
  • the electrodes 435 and 436 are described having a specific polarity, a person of skill in the art will understand that the polarity of the electrodes may be reversed, for example, depending on the biopolymer or macromolecule of interest.
  • the upper assembly 430 can include a sealing element 438 configured to contact a perimeter or boundary of the concave portion 410 of the cartridge 400.
  • the sealing element 438 may be a gasket, an O-ring, or other suitable structure. It is advantageous to provide a positive seal between the upper assembly 430 and the lip 415 to prevent evaporation of a sample from the sample wells 420a and 420b during the electrophoresis process. The evaporation of even a small amount of the sample, or of a liquid contained within the sample may have negative effects on the transport of biopolymers or macromolecules through the nanochannels 428.
  • FIG. 4B depicts a cutaway view of an embodiment of a portion of a system and a cartridge used for nanoanalysis.
  • the substrate 422 may include electrode portions 437 and 438.
  • the electrode portions 437 and 438 are formed in or through the substrate and provide a conducting path from a voltage or current source external to the cartridge to the sample wells 420a and 420b.
  • the electrode portions 437 and 438 may be conducting electrodes embedded in the glass substrate, formed integrally with the substrate, or may be located in through holes in the substrate. At least a portion, for example, a top surface, of the electrode portions 437 and 438 is configured to be in contact with a fluid or liquid sample which is placed in the sample wells 420a.
  • the electrode portions 437 and 438 can create an electric field in the sample wells 420a and 420b sufficient to electrophoretically drive or migrate molecules, such as DNA molecules, from one sample well 420a to an adjacent sample well 420b through the nanostructure wall 426.
  • the electric field can be applied to the sample wells 420a and 420b using a voltage or current source located in the platform 340, or from another location below the cartridge 400.
  • the platform 340 can have electrode pads (not shown) which are disposed in the recess 342 and are aligned with the electrode portions 437 and 438 such that when the cartridge 400 is inserted into the recess 342, electrical contact is established between the electrode pads and the electrode portions 437 and 438.
  • the sample wells 420a and 420b may be arranged in various patterns or networks.
  • a single sample well 420a may be associated with more than one electrodes.
  • the chip 403 may have a single sample well 420a adjacent to three sample wells 420b, separated by a nanostructure wall 426 having regions of nanochannels 428, as described elsewhere herein.
  • Three or more electrodes 435 or electrode portions 437 may be associated with the single sample well 420a, being positioned such that one electrode 435 or electrode portion 437 corresponds to one of the three adjacent sample wells 420b.
  • the chip 403 may have three sample wells 420a adjacent to a single sample well 420b as described elsewhere herein.
  • a single electrode 435 or electrode portion 437 may be associated with each of the sample wells 420a, and three or more electrodes 436 or electrode portions 438 may be associated with the single sample well 420b, being positioned such that one electrode 436 or electrode portion 438 corresponds to one of the three adjacent sample wells 420b.
  • the chip 403 comprises a single sample well 420a or 420b
  • a single electrode 435, 436 or electrode portion 437, 438 may be associated with the single sample well 420a or 420b.
  • the arrangement of electrodes 435, 436, electrode portions 437, 438 and sample wells 420a, 420b are exemplary only, and the scope of the present disclosure is not limited thereto.
  • the well structure 424 receives a small volume of sample (e.g., a 6 ⁇ sample).
  • the process of nanoanalysis and imaging the DNA or biopolymers in the sample may take a long period of time (e.g., up to 24 hours). Over the 24 hours, the small volume sample may partially or completely evaporate to a point where the sample is no longer usable for nanoanalysis and imaging.
  • the volume of the sample is not particularly limited. In some embodiments, the sample size may be less than about 1 ⁇ ,, about 1 ⁇ , about 2 ⁇ ,, about 3 about 8 ⁇ , about 9 ⁇ , about 10 about 15 ⁇ ,, about 20 ⁇ ,, greater than about 20 ⁇ ,, or any other volume there between.
  • the process of nanoanalysis and/or imaging can take less than about 2 hours, about 2 hours, about 4 hours, about 6 hours, about 8 hours, about 10 hours, about 12 hours, about 14 hours, about 16 hours, about 18 hours, about 20 hours, about 22 hours, about 24 hours, about 30 hours, about 36 hours, about 48 hours, greater than about 48 hours, or any amount of time therebetween.
  • the sample can be added to both the sample wells 420a and 420b, as when the driving force is electrophoresis.
  • the sample is added to only sample well 420a, and the sample is moved to sample well 420b via a driving force, such as hydraulic pressure.
  • the cartridge 400 may include a well insert.
  • FIG. 4C depicts an embodiment of a cutaway view of a portion of the cartridge 400.
  • the addition of a well insert to the cartridge 400 decreases the volume of the sample compartment, and can minimize or prevent evaporation or condensation of the sample in the sample wells 420a and 420b.
  • Cartridge 400 includes sample well walls 421 , which extend vertically from well structures 424, generally perpendicular to the planar surface of substrate 422.
  • the tops of the sample well walls 421 include a seal 423 which provides a seal between the sample well walls 421 and the upper assembly 430.
  • the seal is water tight or air-tight, to prevent escape of the sample 425 from the well structures 424.
  • the sample well walls 421 surround each sample well 420a and 420b individually, and seal against the upper assembly via the seals 423.
  • the seals 423 may be constructed similar to the sealing element 438 described above.
  • a small amount of mineral oil 426 can be added to well compartments 424 after the addition of sample 425.
  • the mineral oil 426 provides a vapor seal which acts as a barrier to vaporization or evaporation of the sample 425.
  • FIG. 5 depicts an embodiment of the nanochannels used for nanoanalysis.
  • the components of an embodiment of a nanostructure wall 526 are depicted.
  • the nanostructure wall 526 is adjacent to a sample well 520.
  • the sample well may be filled with a liquid sample containing a biopolymer or macromolecule.
  • the movement of DNA molecules through the nanostructure wall 526 is described herein as an example, and embodiments of the present disclosure are not limited thereto.
  • the nanostructure wall 526 may be divided into two zones: a transition zone 550a and a nanochannel zone 550b.
  • the transition zone 550a includes a lip region 551, one or more feeder channels 553, a pillar region 554, and one or more relaxation channels 557.
  • the lip region 551 is adjacent to a sample well 520 and is a raised portion with respect to the sample well 520.
  • the lip region is the first part of the nanostructure wall 526 that the DNA molecule encounters when being moved, translocated, or otherwise driven from one sample well 520 to another using, for example, electrophoresis.
  • the lip region 5 1 provides an transition area for DNA molecules leaving the sample well 520 and entering the subsequent regions of the nanostructure wall 526.
  • a coiled or entangled DNA molecule 552 is depicted in the lip region 551 , having been driven from the sample well 520.
  • the lip region 551 may have a depth of from about 0.1 microns to about 10 microns, as measured from a top surface of the well structure 524.
  • the lip region may be from about 0.5 micron to about 1000 microns in length, wherein length is defined as being in the direction transversing the nanostructure wall 526 from one sample well 520 to another. In some embodiments, the lip region is about 1.5 microns deep and about 15 microns in length.
  • the dimensions provided herein are exemplary only, and the dimension may be construed to be any value within the listed ranges.
  • the feeder channels 553 Adjacent to the lip region 551 are the one or more feeder channels 553.
  • the feeder channels 553 funnel or direct the coiled or entangled DNA molecules 552 into the pillar region 554.
  • the one or more feeder channels 553 run parallel to each other, and are wide channels, relative to the nanochannels 528.
  • the feeder channels 553 and may have a width of about 0.05 microns to about 25 microns, or any value therebetween, wherein width is understood to be in a direction perpendicular to length as described above.
  • the feeder channels 553 may have a depth of from about 20 nm to about 1000 nm, or any value therebetween. In some embodiments, the feeder channel is about 50 nm in depth and about 1.5 microns wide.
  • the feeder channels 553 lead to the pillar region 554.
  • the pillar region 554 includes a floor 556 which, in some embodiments, is contiguous with the bottom surface of the feeder channels 553.
  • the pillar region 554 also includes or more pillars 555.
  • the pillars 555 may be silicon formations which are interspersed throughout the pillar region, with the pillars 555 extending from the floor 556 of the pillar region to a top portion which is raised above the floor 556.
  • the top portion of the pillar region is in the same plane as the top surface of the well structure 524, and may be in contact with the substrate (not shown).
  • the pillars 555 may be of any shape, that is, the pillars may have a cross-sectional shape which is round, square, diamond, ovoid, rectangular, or any other desired shape.
  • the pillars 555 may vary from one to another in size, shape, height, and distance from other pillars 555.
  • the pillars 555 may be evenly spaced unevenly spaced throughout the pillar region 554.
  • the pillar region 554 may include two zones of pillars 555, wherein the first zone comprises pillars of one a first dimension, shape, and/or height, and the second zone of pillars comprises pillars 555 of a second dimension, shape, and/or height, different from the first dimension.
  • the pillars 555 within the pillar region 554 are sized, shaped, and positioned to untangle, uncoil, or otherwise straighten tangled or coiled biopolymers or macromolecules.
  • the size of the pillars 555 and the spacing between the pillars creates a tortuous flow path through which the coiled or tangled DNA molecule 552 cannot fit.
  • a motive force such as an electric field
  • the coiled or tangled DNA molecule 552 is mechanically forced to uncoil as the molecule interacts with the pillars 555.
  • the spacing between the pillars 555 of the first zone is larger than the spacing of the pillars 555 of the second zone.
  • the first zone causes an initial untangling or uncoiling, before the molecules reach the second zone.
  • the molecules are forced through narrower spaces, which causes a further untangling or uncoiling of the molecules.
  • the distance between pillars can vary.
  • the distance between two pillars can be about 25 nm, about 50 nm, about 100 nm, about 150 nm, about 200 nm, about 250 nm, about 300 nm, about 350 nm, about 400 nm, about 450 nm, about 500 nm, about 1000 nm, about 2000 nm, about 3000 nm, about 4000 nm, about 5000 nm, or a range between any two of these values.
  • the distance between pillars is about 0.1 micron to about 2.5 microns.
  • the pillars may have a height, that is, a distance from the floor 556 to their top surfaces of from about 20 nm to about 5000 nm, or any value therebetween.
  • the pillars may have a width, diameter, or long dimension, depending on their shape, of from about 50 nm to about 10000 nm, or any value therebetween.
  • the pillars 555 have a height of about 50 nm and a width, diameter, or long dimension, of from about 200 nm to about 5000 nm.
  • the pillar region 554 adjoins a plurality of relaxation channels 557.
  • the relaxation channels 557 are channels that act as inlets to the plurality of nanochannels 528.
  • the relaxation channels 557 are funnel shaped channels.
  • the relaxation channels 557 have a wider dimension an end adjacent to the pillar region 554 and a narrower dimension at an end proximate to the nanochannels 528.
  • the relaxation channels 557 receive uncoiled and untangled or partially uncoiled and untangled molecules and help to further linearize the molecules as the molecules enter the plurality of nanochannels 528.
  • a linearized DNA molecule 558 is depicted entering one nanochannel 528 from the associated relaxation channel 557.
  • the relaxation channels 557 may be from about 10 to about 5000 microns long, about 20 nm to 300 nm deep, and about 50-1000 nm wide. In some embodiments, the relaxation channels 557 may be about 80 microns long, 50 nm deep, and 300 nm wide, at their widest point.
  • the plurality of nanochannels 528 receive the linearized DNA molecules, and are sized such that only linearized molecules can fit into and can be transported or moved through the nanochannels 528.
  • the nanochannels 528 may be from about 20 nm to about 300 nm wide, about 30 to about 300 nm deep, and from about 10 to about 10000 microns long. In some embodiments, the nanochannels are about 45 nm wide, about 45 nm deep, and about 350 microns long.
  • FIG. 6 depicts a system for nanoanalysis using a cartridge as described herein. Specifically, a system 600 is used for optical analysis of biopolymers or macromolecules as they move through a plurality of nanochannels. In some embodiments, the biopolymer or macromolecule has been tagged, stained, or marked in order to enable optically imaging of the biopolymer or macromolecule. In some embodiments, DNA may be advantageously marked with fluorescent markers and imaged using the system 600.
  • the system 600 includes an upper assembly 630, and a platform 640, similar to those described elsewhere herein.
  • the cartridge 100 may be received into a receiving portion of the of the platform 640.
  • the system 600 also includes illumination sources 660, illumination optics 670, imaging optics 680, and an imaging device 690.
  • the illumination sources 660 may be lasers, visible light sources, sources of infrared light, sources of ultraviolet light, or any combination thereof suitable to excite a fluorescent tag or other marker.
  • the illumination sources may be 3 lasers, whose wavelengths are 473 nm, 532 nm, and 635 nm, respectively.
  • the illumination optics 670 may include a series of reflective and/or refractive elements, focusing elements, and filtering elements to image the biopolymers or macromolecules which are moving through the nanochannels 528.
  • the imaging optics 680 are disposed generally below the level of the platform 640, and are configured to take an image or picture of the biopolymers and/or macromolecules in the nanochannels 528.
  • the substrate 122 on the cartridge 100 faces downward when recess in the platform 640, and is transparent or translucent.
  • the transparency of the substrate 122 allows the illumination beam to illuminate linearized molecules 558 passing through the nanochannels 528.
  • the transparent substrate 122 also allows for imaging of the linearized, illuminated molecules as they pass through the nanochannels 528.
  • An imaging device 690 such as a camera, a CCD array, infrared or UV sensor, or other imaging device, is connected to the imaging optics 680.
  • the imaging device 690 provides an output corresponding to the imaged linearized molecules detected in the imaging device 690 which can be used in further analysis.
  • platform 640 is moveably mounted.
  • the platform 640 may be connected to a system of translation motors which are capable of moving the platform, and accordingly, the cartridge 100, in the x-y plane.
  • the upper assembly 630 is moveably mounted, and is configured to move the cartridge in the x-y plane. In this way, moving the upper assembly 630 causes movement of the cartridge 100 via the interaction between the upper assembly 630 and the lip of the concave portion of the cartridge.
  • both the platform 640 and the upper assembly 630 are configured to move the cartridge 100 in the x-y direction. [0083] Movement of the platform 640 and/or the upper assembly may be advantageous for imaging the plurality of nanochannels 528.
  • the adjacent nanochannels 528 may not be illuminated, and thus, the imaging device 690, may not be able to image linearized molecules in adjacent nanochannels 528.
  • the imaging optics 680 and imaging device are configured to image only a very small area, for example, a portion of a single nanochannel 528. It may be desirable to only image a small portion of a nanochannel 528 or of several nanochannels 528 in order to produce a high quality image.
  • the platform 640 and/or the upper assembly 630 can move to bring a different portion of the same nanochannels 528, or an adjacent nanochannel 528 into the field of view of the imaging device 690.
  • FIG. 7 depicts an embodiment of a control system for a nanoanalysis system and apparatus.
  • a control system 700 includes a controller 762, a memory 764 and an image processing system 766.
  • the controller 762 is in communication with the memory 764 and the image processing system 766.
  • the controller comprises a processor and an internal memory or cache.
  • the memory 764 may contain computer-readable instructions for operating the controller 762 and/or the control system 700.
  • the memory 764 can store image data received from an imaging device 790.
  • the image processing module 766 may comprise a processor and a memory, and may be configured to receive the image data from the memory 764 or the imaging device 790, and to analyze, evaluate, read, interpret, or otherwise process the image data.
  • the imaging processing module 766 may be configured to evaluate the molecules moving through the nanochannels 428 and make determinations about the identity of the molecules, of specific gene sequences tagged or marked on the molecules, or to perform a variety of other processes to evaluate or identify the molecules imaged in the nanochannels 428.
  • the controller 762 is also in communication with the illumination sources 760, the illumination optics 770, the imaging optics 780, the imaging device 790, one or more x- y translation motors 792, and a motive force generator 794.
  • the controller 762 is configured to power on or off the illumination sources 760 and control the intensity of the illumination sources 760.
  • the controller 762 may be configured to control the direction, focus, or filtering of the illumination beam by controlling the illumination optics 770.
  • the controller 762 is configured to control the imaging optics 780, for example, by controlling the focus, filtering, field of view, or any other desired parameter.
  • the controller 762 sends and receives signals to and from the imaging device 790.
  • the controller 762 may control capture of images, shutter, depth of field, focus, f-stop, exposure time, shutter speed, ISO, whiteness level, brightness, contrast, or adjust any other parameter associated with the imaging device 790.
  • the controller 762 is configured to send control signals to one or more x-y translation motors 792, such as are described herein.
  • the controller 762 may be configured to control operation of an x-y translation motor 792 in the platform 340 in order to move the cartridge 300 to bring portions of the nanochannels into view of the imaging device 790, as needed or desired.
  • the controller 762 can be configured to operate or supply control signals to the motive force generator 794.
  • the motive force generator can be similar to the electrodes or pressure generating elements described herein.
  • the controller 762 can be configured to communicate with a temperature controller 795.
  • the temperature controller 795 is configured to control the temperature control device to maintain the sample at a desired temperature using temperature inputs from the reservoir or platform, and environmental measures, such as relative humidity, ambient temperature, ambient pressure, and the like.
  • the controller 762 operates the imaging system by controlling and coordinating the timing of operating the illumination sources 760, the motive force generator 794, the imaging device 790, and the other portions of the control system 700.
  • the controller can supply a signal to the motive force generator 794 to induce movement of biopolymers or macromolecules in the cartridge as described herein.
  • the controller 762 may remove the signal, or may provide an interrupt signal to stop application of the motive force from the motive force generator 794.
  • the controller 762 may provide a signal to the illumination sources 760 to illuminate a portion of the nanochannels which may have linearized biopolymers or macromolecules within.
  • the controller 762 can signal the imaging optics and the imaging device 790 to image a portion of the nanochannels. After an image of a portion of the nanochannels is obtained, the controller 762 may signal the illumination sources 760 to turn off, and signal the x-y translation motor 792 to move the cartridge a specified amount. After the cartridge has been moved, the controller 762 may re-energize the illumination sources 760 and signal the imaging device 790 to obtain another image. The process may repeat as many times as needed to obtain an image of all the nanochannels, or as desired. This process will be described in more detail with respect to FIG. 8.
  • FIG. 8 is a flow diagram of a process for imaging linearized molecules as they pass through nanochannels.
  • a process 800 for imaging molecules in nanochannels begins in step 802, wherein a sample containing a marked, tagged, or stained biopolymer, such as DNA is added to a first reservoir, or a sample well 420a.
  • a buffer solution or other liquid or fluid may be added to a second reservoir, or a sample well 420b, in order to facilitate the electrophoresis of the biopolymer or macromolecule.
  • a portion of the upper assembly 430 is brought into contact with the cartridge 400, forming a seal around the concave area 410 to prevent evaporation of the sample.
  • the upper assembly includes negative and positive electrodes 435 and 436.
  • the negative electrodes 435 are brought into contact with the sample in the first reservoir, or sample well 420a
  • the positive electrode 436 may be brought into contact with the buffer solution or liquid in the second reservoir, or sample well 420b.
  • the electrode portions 437 and 438 of the substrate 422 are brought into contact with the sample and the buffer solution.
  • the upper assembly 430 has a pressure generation element as described above, which is brought into position as the upper assembly 430 is brought into contact with the cartridge 400.
  • step 804 the motive force is applied to the sample wells 420a and 420b.
  • this is accomplished by applying an electric field to the sample wells by using, for example, the negative and positive electrodes 435 and 436, and/or the electrode portions 436 and 437 of the substrate.
  • this is accomplished by applying a pressure gradient across the nanostructure wall 426 sufficient to drive molecules from the first reservoir to the second reservoir through the nanochannels 428.
  • the process 800 moves to step 806, wherein the motive force is removed after a predetermined amount of time.
  • the predetermined amount of time may be determined based on the biopolymer or macromolecule of interest. In some embodiments, the predetermined amount of time may be determined based on the imaging apparatus, for example, based on the size of the field of view of the imaging apparatus.
  • the time the motive force is applied may be 1 microsecond, 5 microseconds, 10 microseconds, 20 microseconds, 50 microseconds, 0.1 seconds, 0.5 seconds, 1 second, 2 seconds, 3 seconds, 4 seconds, 5 seconds, 10 seconds, 15 seconds, 20 seconds, 25 seconds, 30 seconds, 45 seconds, 60 seconds, 90 seconds, 120 seconds, 5 minutes, 10 minutes, 15 minutes, 20 minutes, or more, or any amount of time therebetween.
  • the process 800 moves to step 808, wherein at least a portion of a predetermined region of the nanochannels is imaged through the transparent substrate 422 or viewing window on the underside of the cartridge 400.
  • the imaging of the portion of the predetermined region uses the illumination sources and imaging device as described herein.
  • the first portion of the nanochannels 428 imaged may include the plurality of nanochannels between a first pair of sample wells 420a and 420b.
  • the first portion imaged may include a single nanochannel 428, or may include only subset of nanochannels 428 between the first pair of sample wells 420a and 420b. It will be understood that the portion imaged may be varied without departing from the scope of the present disclosure.
  • the process 800 moves to step 810 wherein the cartridge 400 is moved so as to bring another portion of the nanochannels 428, different from the previously imaged portion, into position for imaging.
  • the upper assembly 330 or platform 340 may be moved to bring the nanochannels 428 into position for imaging.
  • the illumination optics may be operated to direct the illumination beam at a different portion of the nanochannels 428, which can then be imaged.
  • the second portion imaged may include the plurality of nanochannels between a second pair of sample wells 420a and 420b.
  • the second portion imaged may include another single nanochannel 428 or subset of nanochannels between the first pair of sample wells 420a and 420b, other than those previously imaged.
  • a person of skill in the art will understand that the second portion of the nanochannels 428 imaged may vary without departing from the scope of this application.
  • the process 800 moves to decision state 812 wherein it is determined if each portion of the predetermined region of the nanochannels has been imaged.
  • the region to be imaged can include a specified length of each of the nanochannels 428 running between each pair of sample wells 420a and 420b of cartridge 400. If the predetermined region has not been fully imaged, the process returns to step 810, wherein the imaging of portions of the region of nanochannels continues until the entire predetermined region has been imaged.
  • the process 800 moves to decision state 814, wherein it is determined whether the entire sample has been imaged.
  • the entire sample is imaged after a substantial portion of, or a specific amount of all the molecules of interest inserted into sample compartment have been passed through the nanochannels 428.
  • the specific amount of all the molecules of interest which should pass through the nanochannels 428 may be determined based on the type of molecule being imaged, the volume of the sample wells, or any other desired parameter.
  • the a controller may determine when a sufficient portion of the sample has been imaged based on interpretation of the imaging data. Thus, if the controller has obtained imaging information of a sufficient amount of the molecules of interest, the controller may determine that no further imaging of the sample is needed.
  • step 804 the motive force is applied to the sample wells 420a and 420b, linearized molecules are moved into and/or through the nanochannels 428, and the motive force is removed after the predetermined time has elapsed. This process continues until the entire sample has been imaged, or it is determined that no further imaging is necessary, and the process 800 ends in step 716.
  • steps of process 800 need not be performed in the order specified. Furthermore, a person of skill in the art will understand that the processes may be performed in parallel, and no steps in one process necessarily preclude the performance of steps in another process. In some embodiments, the processes occur in an overlapping fashion, with steps from one process giving rise to or, initiating steps from another process, or steps from one process being triggered by steps from another process.
  • the technology is operational with numerous other general purpose or special purpose computing system environments or configurations.
  • Examples of well-known computing systems, environments, and/or configurations that may be suitable for use with the invention include, but are not limited to, personal computers, server computers, hand-held or laptop devices, multiprocessor systems, processor-based systems, programmable consumer electronics, network PCs, minicomputers, controllers, microcontrollers, mainframe computers, distributed computing environments that include any of the above systems or devices, and the like.
  • instructions refer to computer-implemented steps for processing information in the system. Instructions can be implemented in software, firmware or hardware and include any type of programmed step undertaken by components of the system.
  • a processor may be any conventional general purpose single- or multi-chip processor such as a Pentium ® processor, a Core 13, 15, or 17 processor, a 8051 processor, an AMD FX series processor, a MIPS ® processor, an Atom processor, an Alpha ® processor, or any other desired or suitable processor.
  • the processor may be any conventional special purpose processor such as a digital signal processor or a graphics processor.
  • the processor typically has conventional address lines, conventional data lines, and one or more conventional control lines.
  • each of the modules comprises various subroutines, procedures, definitional statements and macros.
  • Each of the modules are typically separately compiled and linked into a single executable program. Therefore, the description of each of the modules is used for convenience to describe the functionality of the preferred system.
  • the processes that are undergone by each of the modules may be arbitrarily redistributed to one of the other modules, combined together in a single module, or made available in, for example, a shareable dynamic link library.
  • the system may be used in connection with various operating systems such as Linux®, UNIX® or Microsoft Windows®.
  • the system may be written in any conventional programming language such as C, C++, BASIC, Pascal, or Java, and run under a conventional operating system.
  • C, C++, BASIC, Pascal, Java, and FORTRAN are industry standard programming languages for which many commercial compilers can be used to create executable code.
  • the system may also be written using interpreted languages such as Perl, Python or Ruby.
  • DSP digital signal processor
  • ASIC application specific integrated circuit
  • FPGA field programmable gate array
  • a general purpose processor may be a microprocessor, but in the alternative, the processor may be any conventional processor, controller, microcontroller, or state machine.
  • a processor may also be implemented as a combination of computing devices, e.g., a combination of a DSP and a microprocessor, a plurality of microprocessors, one or more microprocessors in conjunction with a DSP core, or any other such configuration.
  • the functions and methods described may be implemented in hardware, software, or firmware executed on a processor, or any combination thereof. If implemented in software, the functions may be stored on or transmitted over as one or more instructions or code on a computer-readable medium.
  • Computer-readable media include both computer storage media and communication media including any medium that facilitates transfer of a computer program from one place to another.
  • a storage medium may be any available media that can be accessed by a computer.
  • such computer-readable media can comprise RAM, ROM, EEPROM, CD-ROM or other optical disk storage, magnetic disk storage or other magnetic storage devices, or any other medium that can be used to carry or store desired program code in the form of instructions or data structures and that can be accessed by a computer.
  • any connection is properly termed a computer-readable medium.
  • the software is transmitted from a website, server, or other remote source using a coaxial cable, fiber optic cable, twisted pair, digital subscriber line (DSL), or wireless technologies such as infrared, radio, and microwave
  • the coaxial cable, fiber optic cable, twisted pair, DSL, or wireless technologies such as infrared, radio, and microwave are included in the definition of medium.
  • Disk and disc includes compact disc (CD), laser disc, optical disc, digital versatile disc (DVD), floppy disk and Blu-ray disc where disks usually reproduce data magnetically, while discs reproduce data optically with lasers. Combinations of the above should also be included within the scope of computer-readable media.

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Abstract

La présente invention concerne des cartouches et des systèmes de nanoanalyse et des procédés d'utilisation des cartouches et des systèmes Par exemple, une cartouche peut comprendre une puce formant un ou plusieurs puits d'échantillon avec une pluralité de nanotubes s'étendant entre une paire de puits d'échantillon adjacents, les nanotubes étant conçus pour recevoir des molécules linéarisées. Lors de la linéarisation, les molécules dans les nano-canaux sont imagées par le biais d'un substrat transparent sur lequel la puce est située. Le mouvement des molécules à travers les nano-canaux peut être démarré et stoppé et la cartouche peut être déplacée pour faciliter l'imagerie des molécules.
PCT/US2014/049442 2013-08-02 2014-08-01 Système pour nanoanalyse Ceased WO2015017801A1 (fr)

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WO2016196625A1 (fr) * 2015-06-02 2016-12-08 Nanopore Diagnostics, Llc Détection d'acides nucléiques
DE102015218422A1 (de) 2015-09-24 2017-03-30 Universität Stuttgart Sensorelement für Photolumineszenz-Messungen, Photolumineszenz-Detektionseinrichtung, Verfahren zum Betreiben einer Photolumineszenz-Detektionseinrichtung, Verfahren zur Herstellung eines Sensorelements, und Verwendung eines Sensorelements
WO2024145269A1 (fr) 2022-12-25 2024-07-04 Bionano Genomics, Inc. Système de cartographie de génome optique
WO2024145268A2 (fr) 2022-12-25 2024-07-04 Bionano Genomics, Inc. Cartouches de mappage de génome optique
WO2024145270A1 (fr) 2022-12-25 2024-07-04 Bionano Genomics, Inc. Instrumentation de systèmes de cartographie de génome optique

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WO2016196625A1 (fr) * 2015-06-02 2016-12-08 Nanopore Diagnostics, Llc Détection d'acides nucléiques
DE102015218422A1 (de) 2015-09-24 2017-03-30 Universität Stuttgart Sensorelement für Photolumineszenz-Messungen, Photolumineszenz-Detektionseinrichtung, Verfahren zum Betreiben einer Photolumineszenz-Detektionseinrichtung, Verfahren zur Herstellung eines Sensorelements, und Verwendung eines Sensorelements
WO2024145269A1 (fr) 2022-12-25 2024-07-04 Bionano Genomics, Inc. Système de cartographie de génome optique
WO2024145268A2 (fr) 2022-12-25 2024-07-04 Bionano Genomics, Inc. Cartouches de mappage de génome optique
WO2024145270A1 (fr) 2022-12-25 2024-07-04 Bionano Genomics, Inc. Instrumentation de systèmes de cartographie de génome optique

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