WO2022098064A1 - Micropuce et son procédé de scellement - Google Patents

Micropuce et son procédé de scellement Download PDF

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Publication number
WO2022098064A1
WO2022098064A1 PCT/KR2021/015738 KR2021015738W WO2022098064A1 WO 2022098064 A1 WO2022098064 A1 WO 2022098064A1 KR 2021015738 W KR2021015738 W KR 2021015738W WO 2022098064 A1 WO2022098064 A1 WO 2022098064A1
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WIPO (PCT)
Prior art keywords
reaction
plate
microchip
channels
fluid
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Application number
PCT/KR2021/015738
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English (en)
Korean (ko)
Inventor
변재영
장희영
박현규
Original Assignee
주식회사 미코바이오메드
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Publication of WO2022098064A1 publication Critical patent/WO2022098064A1/fr

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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
    • B01L7/00—Heating or cooling apparatus; Heat insulating devices
    • B01L7/52—Heating or cooling apparatus; Heat insulating devices with provision for submitting samples to a predetermined sequence of different temperatures, e.g. for treating nucleic acid samples
    • 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
    • B01L2200/0684—Venting, avoiding backpressure, avoid gas bubbles
    • 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
    • B01L2300/0861—Configuration of multiple channels and/or chambers in a single devices
    • 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
    • B01L2300/16—Surface properties and coatings
    • 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
    • B01L2400/08—Regulating or influencing the flow resistance
    • B01L2400/084—Passive control of flow resistance
    • B01L2400/086—Passive control of flow resistance using baffles or other fixed flow obstructions

Definitions

  • the present invention relates to a microchip and a sealing method thereof.
  • the microchip has the ability to simultaneously perform various experimental conditions by flowing a fluid through the microfluidic channel.
  • a microchannel is made using a substrate (or chip material) such as plastic, glass, or silicon, a fluid (eg, a liquid sample) is moved through the channel, and then mixed in a plurality of chambers in the microchip. and react.
  • the microchip is also called "lab-on-a-chip" in that experiments conventionally performed in a laboratory are performed in a small chip.
  • Microchips have created cost and time savings in various fields such as pharmaceuticals, biotechnology, medicine, and chemistry, and can provide high accuracy, efficiency, and reliability. For example, by using a microchip, the amount of expensive reagents used for cell culture, proliferation, and differentiation can be significantly reduced compared to the conventional method, thereby significantly reducing costs. In addition, since a protein sample or cell sample is used in a much smaller amount than the conventional method and image analysis is possible using the same, the amount of use or consumption of the sample and the analysis time can be reduced.
  • the sealing stopper technology such as rubber and silicone sealing is a technology that configures the stopper to match the shape of the inlet and outlet of the fluid, and presses it with a physical support to prevent leakage.
  • the sealing stopper technology has the inconvenience of having to change the shape of the stopper according to the shape of the inlet and the outlet.
  • the sealing stopper technology requires an additional physical support to prevent thermal expansion of the reaction fluid, and a driving unit for controlling this is required, so there is a problem in that the unit price is increased.
  • An object of the present invention is to solve the above problems, and to provide a microchip, a method for sealing the microchip, and an integrated chip including the microchip.
  • An embodiment of the present invention provides an inlet through which a fluid is introduced, one or more reaction channels in which a predetermined reaction is performed with respect to the introduced fluid, and an outlet through which the fluid flows from the reaction channel, and the inlet or outlet is porous therein sintered polymer layer; It provides a microchip, characterized in that the porous sintered polymer layer swells when in contact with water to seal the inlet or outlet.
  • the polymer is ultra-high molecular weight polyethylene (UHMW-PE), hydroxypropylmethylcellulose, hydroxyethylmethylcellulose, hydroxypropylcellulose, hydroxymethylcellulose, hydroxyethylcellulose, carboxymethylcellulose, Calcium carboxymethylcellulose, sodium carboxymethylcellulose, methylcellulose, ethylcellulose, polyethylene oxide, leukostbin gum, guar gum, xanthan gum, acacia gum, tragacanth gum, alginic acid, sodium alginate , calcium alginate, ammonium alginate, agar, gelatin, poloxamer, polymethmethylacrylate, carbomer, polycarbophil, polyvinylpyrrolidone, polyvinylacetate, polyethylene glycol, polyvinylpyrrolidone-poly Vinyl acrylate copolymer, polyvinyl alcohol-polyethylene glycol copolymer, polyvinylpyrrolidone-polyvinyl acetate
  • UHMW-PE
  • the polymer layer may be provided at the same or higher position than the upper position of the reaction channel.
  • the polymer may be a bead having a diameter of 10 ⁇ m to 800 ⁇ m, or an aggregate of beads having a diameter of 5 mm or less.
  • a barrier layer may be further provided on the polymer layer to be formed in multiple stages.
  • the inlet and/or outlet may further include a barrier layer provided on top of the polymer layer, and the barrier layer may block the movement of nucleic acids.
  • the inlet and/or outlet may further include a polymer layer on top of the barrier layer.
  • the barrier layer may include polyethylene.
  • the microchip is a flat plate-shaped first plate, a second plate vertically disposed on the first plate to form a reaction channel, and an upper portion of the second plate spaced apart from the first plate It may include a third plate disposed on the substrate having an inlet and an outlet formed through the substrate.
  • the microchip includes: a lower plate recessed from the surface of a plate-shaped substrate to form a reaction channel; and an upper plate disposed on the lower plate and having an inlet and an outlet formed through the substrate; may include
  • the reaction channel may include a reaction region located inside the outlet and spaced apart.
  • the microchip may further include a plurality of dummy regions, and a plurality of reaction regions may be arranged side by side between the dummy regions.
  • the microchip may be a PCR reaction chip in which a nucleic acid amplification reaction occurs.
  • the microchip and a plurality of microfluidic channels and a pre-treatment chip including a valve provided between the channels and the channels; It provides an integrated chip comprising a, characterized in that the reaction channel and the microfluidic channel is connected.
  • the microchip is a PCR reaction chip having a plurality of reaction channels in which a nucleic acid amplification reaction occurs, and the pretreatment chip includes a plurality of microfluidic channels and a valve provided between the channels and the channels. It may be a nucleic acid pretreatment chip.
  • One embodiment of the present invention is provided with a porous sintered polymer layer for sealing the inlet or outlet by swelling when in contact with water inside the inlet through which the fluid flows or the outlet through which the fluid flows.
  • the microchip includes: an inlet through which a fluid is introduced; an outlet through which the fluid flows; and one or more reaction channels through which a predetermined reaction is performed with respect to the introduced fluid.
  • the polymer layer may be provided at the same or higher position than the upper position of the reaction channel.
  • the sealing method may further include a barrier layer provided on the polymer layer, wherein the barrier layer may block the movement of nucleic acids.
  • the microchip and the integrated chip can be simply sealed without additional equipment by using the sealing member.
  • the present invention does not additionally require a separate valve or a driving connection part of the chip, the size and structure of the chip can be configured simply.
  • the present invention does not require a separate sealing device and a sealing unit, there is an advantage in that multiple channels can be installed by utilizing the space in the chip.
  • the present invention has an advantage in that the equipment is miniaturized.
  • the sealing effect can be realized quickly, and thus the driving time and cost can be reduced.
  • FIG. 1 is a plan view showing a microchip according to an embodiment of the present invention.
  • FIG. 2 is a cross-sectional view of the inside of a microchip according to an embodiment of the present invention as viewed from the side.
  • 3 and 4 show a top view and a bottom view from below of a PCR chip according to an embodiment of the present invention.
  • FIG. 5 is a cross-sectional view of the interior of an integrated chip according to an embodiment of the present invention viewed from above.
  • FIG. 6 is a bottom view of the interior of the integrated chip according to an embodiment of the present invention as viewed from below.
  • FIG. 7 is a cross-sectional view of the interior of the integrated chip according to the embodiment of the present invention as viewed from the side.
  • FIG. 8 is a perspective view illustrating a first valve according to an embodiment of the present invention.
  • FIG. 9 is a cross-sectional view of a first valve and a microfluidic channel viewed from the side according to an embodiment of the present invention.
  • FIG. 10 is a perspective view illustrating a first valve according to an embodiment of the present invention.
  • 1 and 2 are a plan view and a cross-sectional view of a microchip including a sealing member according to an embodiment of the present invention.
  • 1 and 2 as a microchip capable of controlling the movement of a fluid, an inlet through which a fluid is introduced, a reaction channel through which a predetermined reaction is performed with respect to the introduced fluid, and an outlet through which the fluid flows out of the reaction channel includes wealth.
  • the fluid flowing in through the inlet may undergo a predetermined reaction in the reaction region, and then may flow out through the outlet.
  • the reaction may be a PCR (nucleic acid amplification) reaction, but this is exemplary and various reactions may be performed according to the embodiment to which the present invention is applied. A description of the microchip will be described later.
  • a sealing member may be included in the inlet or outlet to seal the inlet or outlet.
  • the sealing member serves to prevent at least a portion of the fluid from being lost to the outside in the process of performing a predetermined reaction, and to block contact with external air.
  • the reaction channel may be formed in a straight line such that the inlet and the outlet extend in the longitudinal direction.
  • the microchip may be equipped with one or more reaction channels. Two or more reaction channels may exist according to the purpose and scope of use of the microchip according to an embodiment of the present invention, and specifically 4 to 6 reaction channels may exist.
  • the inlet and outlet may be disposed at positions spaced apart from each other so as not to cause interference between the reaction region and the detection material in the reaction channel.
  • the fluorescent material may come out from the sealing member, which may cause interference with the PCR fluorescence value, which may cause a diagnosis error. Therefore, it is preferable that the inlet and the outlet are disposed at a sufficiently distant distance from the area where a predetermined reaction occurs in the reaction channel so that such interference does not occur.
  • microchip 1 shown in FIGS. 1 and 2 is exemplary, and microchips of various shapes or structures may be used according to an embodiment to which the present invention is applied.
  • the microchip may be utilized as various chips requiring sealing, such as a biochip and a diagnostic chip.
  • each of the inlet and outlet may include an opening through which a fluid is introduced and a protrusion protruding adjacent to the opening.
  • a sealing member may be disposed inside the opening surrounded by the protrusion to seal the opening. Accordingly, the sealing member may be provided at a position higher than the upper surface of the reaction channel.
  • the sealing member may be provided at the same position as the upper position of the reaction channel or higher than the upper position of the reaction channel.
  • the sealing member is also applicable to the inlet and/or outlet.
  • the sealing member may be applied to a position where it is necessary to prevent leakage of fluid, and the position of the sealing member is not necessarily limited to this description.
  • the sealing member for sealing the inlet and/or outlet of the reaction channel is made of any one selected from a polymer resin, an amorphous material, and a metal, and in particular, chlorinated polyethylene, ethylene propylene dimethyl, silicone rubber, acrylic resin, amide-based Resin, epoxy resin, phenol resin, polyester resin, polyethylene resin, ethylene-propylene rubber, polyvinyl butyral resin, polyurethane resin, and nitrile- may be formed to include at least one of butadiene rubber, sealing It is sufficient that the member prevents leakage of fluid and blocks external air, and the present description is not limited thereto.
  • the sealing member may be performed according to various techniques known in the art.
  • the sealing member may be a porous sintered polymer layer 80 that seals the inlet and/or outlet by swelling when in contact with water therein.
  • the porous sintered polymer layer may include porous sintered polymer beads or an aggregate of porous sintered polymer beads.
  • the diameter of the bead may be 10 ⁇ m or more to 800 ⁇ m, and the diameter of the bead aggregate may be 5 mm or less.
  • the diameter of the polymer bead and the diameter of the bead aggregate are not limited to the present description.
  • the porous sintered polymer is a sintering water-swelling polymer and has air permeability. As soon as it comes into contact with moisture, the pores are expanded to prevent the fluid from leaking to the outside.
  • the water-swellable polymer may swell upon contact with water to form a gel layer.
  • the swelling refers to a phenomenon in which a high molecular compound absorbs a solvent to increase its volume.
  • the sintering refers to a process in which powder particles become a single mass through a thermal activation process. When powder or a mass of compressed powder is heated to a temperature below the melting point, the powder melts and adheres to each other and solidifies. It is generally applied to the manufacture of ceramics or small plastics.
  • a water-swelling polymer is a polymer that exhibits fluid absorption due to the introduction of a hydrophilic group in a three-dimensional network structure or a single chain structure through crosslinking between polymer chains. It contains at least 15 times the weight of the polymer itself. may contain, capable of supporting a sufficient amount of fluid under a load, and may contain aqueous solutions, but insoluble in aqueous solutions. In addition, when it enters water, it swells instantaneously and becomes gelled.
  • the porous sintered polymer includes ultra-high molecular weight polyethylene (UHMW-PE), hydroxypropylmethylcellulose, hydroxyethylmethylcellulose, hydroxypropylcellulose, hydroxymethylcellulose, hydroxyethylcellulose, carboxymethylcellulose, carboxymethylcellulose calcium, Carboxymethylcellulose sodium, methylcellulose, ethylcellulose, polyethylene oxide, leukostbin gum, guar gum, xanthan gum, acacia gum, tragacanth gum, alginic acid, sodium alginate, calcium alginate, alginic acid Ammonium, agar, gelatin, poloxamer, polymethmethylacrylate, carbomer, polycarbophil, polyvinylpyrrolidone, polyvinyl acetate, polyethylene glycol, polyvinylpyrrolidone-polyvinyl acrylate copolymer , polyvinyl alcohol-polyethylene glycol copolymer, polyvinylpyrrolidone-polyviny
  • the inlet 40 and/or outlet 50 of the microchip 1 may include the porous sintered polymer layer 80 .
  • one embodiment of the present application may include a multi-layer layer further including a barrier layer 90 provided on the porous sintered polymer layer 80 .
  • one embodiment of the present application may include both the polymer layer 80 positioned above the barrier layer 90 and the polymer layer 80 positioned below the barrier layer 90 .
  • the barrier layer 90 may be a pipette tip filter layer, and any layer that blocks the movement of nucleic acids may be applied.
  • it may be made of a material comprising polyethylene.
  • the amplified nucleic acid remaining inside the chip is retained, and when the porous sintered polymer layer recovers air permeability, there is a possibility that it will leak out as an aerosol.
  • the barrier layer 90 is disposed on the porous sintered polymer layer, there is an effect of preventing leakage of nucleic acids to the outside.
  • the sealing member by using the sealing member, the size and structure of the chip can be simplified, and since a separate sealing device and a sealing unit are not required, a plurality of channels can be installed using the remaining space of the chip. can In addition, it is possible to quickly implement the sealing effect without additional equipment, thereby reducing driving time and cost. In addition, there is an advantage in that it is not necessary to precisely control the movement of the fluid on the movement path of the fluid on the chip. When the fluid comes into contact with the sealing member including the polymer layer, the inlet and/or outlet are sealed and the movement of the fluid is blocked, so there is no need to precisely control the flow rate and the moving speed of the fluid. Therefore, it is possible to miniaturize the equipment, and since the price of the equipment is reduced, there is an economic advantage.
  • the microchip 1 is a first plate 10 in the form of a flat plate, a second plate 20 vertically disposed on the upper portion of the first plate 10 to form a reaction channel , It is spaced apart from the first plate 10 and disposed on the second plate 20 to include a third plate 30 having an inlet 40 and an outlet 50 formed through the substrate.
  • the microchip (1) is recessed from the surface of the plate-shaped substrate, the lower plate (10') to form a reaction channel; and an upper plate 30' disposed on the lower plate 10' and having an inlet 40 and an outlet 50 formed through the substrate; may include
  • the first plate 10 or the lower plate 10' is implemented in a flat plate shape, and may serve as a bottom support of the microchip 1 according to an embodiment of the present invention.
  • the first plate 10 may be implemented with various materials, but preferably, polydimethylsiloxane (PDMS), cycloolefin copolymer (cycle olefin copolymer, COC), polymethylmetharcylate , PMMA), polycarbonate (PC), polypropylene carbonate (PPC), polyether sulfone (PES), and polyethylene terephthalate (PET), and combinations thereof.
  • PDMS polydimethylsiloxane
  • COC cycle olefin copolymer
  • COC polymethylmetharcylate
  • PMMA polycarbonate
  • PC polypropylene carbonate
  • PES polyether sulfone
  • PET polyethylene terephthalate
  • It may be a material selected from, and according to an embodiment, at least a portion of the first plate
  • the surface of the first plate 10 may be treated to have a hydrophilic surface.
  • the hydrophilic material may be a variety of materials, but is preferably selected from the group consisting of a carboxyl group (-COOH), an amine group (-NH2), a hydroxyl group (-OH), and a sulfone group (-SH). Treatment of the hydrophilic material may be performed according to a method known in the art.
  • the second plate 20 is disposed on the first plate 10, and may serve to form the reaction channel 60 of the PCR microchip 1 according to an embodiment of the present invention.
  • the second plate 20 may be implemented with various materials, but preferably, polymethylmethacrylate (PMMA), polycarbonate (PC), cycloolefin copolymer (COC), poly Amide (polyamide, PA), polyethylene (PE), polypropylene (polypropylene, PP), polyphenylene ether (PPE), polystyrene (PS), polyoxymethylene (POM), poly Etheretherketone (polyetheretherketone, PEEK), polytetrafluoroethylene (PTFE), polyvinylchloride (PVC), polyvinylidene fluoride (PVDF), polybutyleneterephthalate (PBT) ), fluorinated ethylenepropylene (FEP), perfluoralkoxyalkane (PFA), and may be a thermoplastic or thermosetting resin material selected from the group consisting of
  • the third plate 30 is disposed on the second plate 20 or the upper plate 30 ′ is disposed on the lower plate 10 ′, so as to cover the reaction channel 60 of the microchip 1 . It may serve as a cover that does this, and a bubble removing unit (not shown) may be formed to protrude into the reaction chamber on the lower surface of the third plate 30 or the upper plate 30 ′.
  • the third plate 30 may be implemented with various materials, but preferably, polydimethylsiloxane (PDMS), cycloolefin copolymer (COC), polymethylmetharcylate (PMMA) ), polycarbonate (PC), polypropylene carbonate (PPC), polyether sulfone (PES), and polyethylene terephthalate (PET), and combinations thereof. material, and according to an embodiment, at least a part of the third plate 30 may be implemented with a light-transmitting material.
  • PDMS polydimethylsiloxane
  • COC cycloolefin copolymer
  • PMMA polymethylmetharcylate
  • PC polycarbonate
  • PPC polypropylene carbonate
  • PES polyether sulfone
  • PET polyethylene terephthalate
  • At least some of the inner surfaces of the first plate 10 , the second plate 20 , and the third plate 30 may be subjected to surface treatment.
  • at least some of the inner surfaces of the lower plate 10 ′ and the upper plate 30 ′ may be subjected to surface treatment.
  • a material such as a silane series or bovine serum albumin (BSA). It can be performed according to various techniques known in the art.
  • the microchip 1 may be a PCR reaction chip 500 in which a nucleic acid amplification reaction occurs.
  • the reaction channel 530 may include a reaction region 540 spaced apart from the outlet 550 therein. If one or more, two or more, or 4 to 6 reaction channels are provided, one or more, two or more, or 4 to 6 reaction zones may be provided correspondingly.
  • the reaction region 540 in the reaction channel 530 is spaced apart from the outlet 550 , and the reaction region 540 is preferably located in the middle of the reaction channel 530 .
  • the PCR reaction chip 500 may further include a dummy region 520 .
  • the dummy region may be provided in plurality, and a plurality of reaction regions may be arranged in parallel between the plurality of dummy regions.
  • the reaction areas are arranged side by side between the dummy areas, there is an advantage in that the heat of the reaction areas is uniformly controlled.
  • the temperature of the two reaction areas disposed on both side portions may be lower than that of the reaction area located in the center.
  • the two reaction areas arranged on both sides may have a lower temperature than the reaction area located in the center due to heat transfer due to heating of the surrounding plastic. Therefore, there is a problem that the efficiency of the PCR reaction may be inhibited compared to the reaction region 540 located in the center. Accordingly, since the dummy regions located on the sides of the two reaction regions disposed on both side portions prevent the temperature of the reaction regions located at the edges from dropping, it is possible to maintain a uniform temperature in all the reaction regions.
  • 5 to 7 show a cross-sectional view, a rear view, and a side cross-sectional view of an integrated chip according to an embodiment of the present invention.
  • the microchip and a plurality of microfluidic channels and a pre-treatment chip including a valve provided between the channels and the channels; It includes and provides an integrated chip in which the reaction channel and the microfluidic channel are connected.
  • the microchip is a PCR reaction chip 500 having a plurality of reaction channels in which a nucleic acid amplification reaction occurs
  • the pretreatment chip 200 includes a plurality of microfluidic channels and between the channels and the channels. It may be a nucleic acid pretreatment chip including a provided valve.
  • the integrated chip 100 is a nucleic acid pre-treatment chip 200 including a plurality of microfluidic channels and valves 300 and 400 provided between the channels and the channels. ; and a PCR chip including a plurality of PCR reaction channels 530 arranged to extend in the longitudinal direction of the channel from one surface of the pretreatment chip.
  • the PCR chip may be connected from the side of the pretreatment chip, but is not limited thereto.
  • the integrated chip 100 pre-processes the introduced samples and reagents and distributes them to a plurality of PCR reaction channels 530 to enable multiple diagnosis.
  • the valve includes a first valve 300 and a second valve 400, and the shape of the valve is not limited.
  • the fluid is introduced through the inlet, transferred through the microfluidic channel, and sequentially moved to the PCR reaction channel 530 through the first valve 300 and the second valve 400 .
  • the first valve 300 has an inlet portion corresponding to a first channel 235 to be described later and an outlet portion corresponding to a second channel 237 to be described later on the side
  • the second valve 400 is a second channel ( 237) and an outlet corresponding to the third channel 239, which will be described later, may be provided on the side.
  • the first valve 300 and the second valve 400 may each control opening and closing of the valve by rotating movement.
  • the first valve 300 and the second valve 400 may slide in a vertical or horizontal direction to control opening and closing of the valve, respectively. Accordingly, it is possible to control the flow of the fluid flowing into the chamber in the valve.
  • the shape of the sliding valve is not particularly limited. For example, when the first valve moves horizontally in one direction, both the first channels and the second channel are opened to allow fluid to flow into or out of the chamber, and when the valve moves horizontally in the opposite direction, the second channel is opened. Both the first channels and the second channels may be closed to prevent a fluid from flowing into or out of the chamber.
  • a horizontal sliding driving method of the second valve may be the same as that of the first valve.
  • both the first channels and the second channel are opened to allow fluid to flow into or out of the chamber, and when the valve moves vertically in the opposite direction, the first Both the channels and the second channels may be closed to prevent fluid from flowing into or from the chamber.
  • a vertical sliding driving method of the second valve may be the same as that of the first valve.
  • the first valve 300 and the second valve 400 may elastically move to control opening and closing of the valves, respectively.
  • Springs are inserted into the valves, and the valves can elastically move vertically by pressing the springs.
  • both the first channels and the second channels are opened to allow fluid to flow into or out of the chamber, and when the first valve moves elastically and is compressed, the first channels and All of the second channels may be closed to prevent fluid from flowing into or out of the chamber.
  • the opening and closing of the first valve 300 and the second valve 400 may be controlled by hinge movement using a hinge means. Opening and closing of the valve and opening and closing of the microfluidic channel can be controlled by moving the hinge, and thus the fluid flowing in the microfluidic channel can be controlled.
  • the microfluidic channel has one end connected to the inlet part, and a plurality of first channels for transferring a biological sample or reagent to the first valve 300 or the mixing chamber 315 in the first valve. (235), a second channel 237 and the second valve having one end connected to the first valve 300 and transferring the nucleic acid to the second valve 400 or the distribution chamber 415 in the second valve ( One end is connected to 400) and may include a plurality of PCR reaction channels 530 for transferring nucleic acids and a third channel 239 corresponding to the plurality of PCR reaction channels 530 .
  • the third channel 239 may be, for example, 4 to 6 channels, and the flow rate of the fluid is precisely distributed according to the number of channels, so that the fluid can be transferred to 4 to 6 channels at the same time.
  • the integrated chip is a first plate 210 of a flat plate shape, is disposed on the upper portion of the first plate 210, microfluidic channels (235, 237, 239), chambers (315, 415), valves ( a second plate 230 forming 300 , 400 and a reaction channel 530 ; It is spaced apart from the first plate 210 and is disposed on the second plate 230 and includes a third plate 250 having an inlet portion 253 and an outlet portion 550 formed through the substrate.
  • an adhesive member 290 may be provided between the second plate 230 and the third plate 250 to seal it.
  • the lower plate 210' is recessed from the surface of the substrate to form microfluidic channels (235, 237, 239), chambers (315, 415), or reaction channels (530); and an upper plate 250 having an inlet portion 253 and an outlet portion 550 formed by being depressed or penetrating from the surface of the substrate, and having valves 300 and 400 formed by protruding from the surface of the substrate, and the lower plate 210 ') and an adhesive member 290 for sealing the upper plate 250'.
  • the plurality of inlets 253 may be formed to pass through the third plate or the upper plate 250 .
  • the reagent inlet 253 may be provided in a plurality of numbers so that a plurality of reagents may be respectively introduced through the plurality of inlets.
  • the third plate 250 and the first plate 210 , or the upper plate 250 and the lower plate 210 may be spaced apart from each other, and a microfluidic channel may be formed in a space between the respective substrates.
  • a space between the second plate 230 vertically arranged, and the third plate 250 and the first plate 210 spaced apart up and down may form a microfluidic channel.
  • the first plate 210 or the lower plate 210 is implemented in a flat plate shape, and may serve as a bottom support of the integrated chip 100 according to an embodiment of the present invention.
  • the first plate 210 may be implemented with various materials, but the description of the material of the first plate 10 of the microchip is the same.
  • the second plate 230 is disposed on top of the first plate 210, and serves to form a microfluidic channel and a PCR reaction channel 530 of the PCR integrated chip 100 according to an embodiment of the present invention. can be performed.
  • the second plate 230 may be implemented with various materials, but the description of the material of the second plate 30 of the microchip is the same.
  • the third plate 250 is disposed on the second plate 230, or the upper plate 250 is disposed on the lower plate 210, the microfluidic channel and the PCR reaction channel 530 of the integrated chip 100. It may serve as a cover to cover, and a bubble removing unit (not shown) may be formed to protrude into the reaction chamber on the lower surface of the third plate 250 or the upper plate 250 .
  • the description of the material of the third plate 250 is the same as the description of the material of the third plate 30 of the microchip.
  • At least some of the inner surfaces of the first plate 210 , the second plate 230 , and the third plate 250 may be subjected to surface treatment. there is. It is the same as the description of the first to third plates of the microchip.
  • an adhesive member (not shown) disposed between the third plate 250 and the first plate 210 or between the upper plate 250 ′ and the lower plate 210 ′ is a fluid between the two substrates. It prevents the risk of leakage and has the effect of blocking the outside air.
  • the adhesive member (not shown) may be an acrylic, silicone, or rubber-based member as a member having adhesive properties, but is not particularly limited. When the adhesive member is disposed adjacent to the inlet 253 and the periphery of the microfluidic channel, the sealing force may be further increased.
  • the first valve 300 and the second valve 400 may be disposed in an extension direction along the length of the micro-oil channel to control the flow of the fluid. According to the control of the valve, the sample pretreatment process, the nucleic acid extraction process, and the process of transferring the nucleic acid to the PCR reaction channel may be sequentially performed.
  • the first valve may have an inlet portion corresponding to the first channel and an outlet portion corresponding to the second channel on the side, and opening and closing the first valve by turning movement can be adjusted
  • the second valve differs only in the function of the first valve and the chamber, but the overall structure and appearance are similar, so the drawing may be substituted for the first valve of FIG. 8 .
  • the second valve may have an inlet portion corresponding to the second channel and an outlet portion corresponding to the third channel on the side, and opening and closing of the second valve may be controlled by rotating movement.
  • the second valve 400 serves to distribute the fluid introduced from the first valve 300 to a plurality of PCR reaction channels 530 . Due to the second valve 400, sophisticated nucleic acid distribution to each of the PCR reaction channels 530 is possible, and the number of detectable targets in each channel can be varied, so that multiple diagnosis is possible. For example, if 4 fluorescent labels are used per 1 PCR reaction channel, 4 targets can be detected and diagnosed. In addition, in the case of a conventional PCR chip, at least 20-50 ⁇ l of fluid is introduced per one PCR reaction channel 530, but in the case of the PCR reaction channel according to an embodiment of the present invention, the minimum amount per one PCR reaction channel is 4 ⁇ It is sufficient to introduce only 8 ⁇ l of fluid.
  • the capacity of the fluid may vary. Therefore, it is possible to form a PCR reaction channel 530 having a finer diameter, and thus a plurality of PCR reaction channels 530 can be arranged by utilizing the remaining space. Since the number of detection targets can be detected as much as a value obtained by multiplying the maximum number of channels and the number of fluorescent materials, there is an advantage that multiple diagnostics can be performed simultaneously in a short time. Specifically, when 4 fluorescent materials are used in 6 channels, it is possible to detect 16 or more targets and perform multiple diagnosis within 30 to 50 minutes.
  • the valve is formed to protrude from the surface of the third plate 250 or the upper plate 250, and the opening and closing are controlled according to the turning movement, so that the inlet part 311 of the microfluidic channel. and the outlet 317 may be sealed.
  • the sealing effect may be further enhanced by providing the sealing member 350 inside the valve.
  • the sealing member is made of any one selected from a polymer resin, an amorphous material, and a metal, and in particular, chlorinated polyethylene, ethylene propylene dimethyl, silicone rubber, acrylic resin, amide resin, epoxy resin, phenol resin, polyester resin , polyethylene-based resin, ethylene-propylene rubber, polyvinyl butyral resin, polyurethane resin, and nitrile- may be formed including at least one selected from the group consisting of butadiene-based rubber. Since the valve is used, the process of using a conventional sealing member or applying heat to seal both sides can be omitted, so that the sealing process can be simplified and the sealing effect is better. Moreover, the effect is better because it is double-sealed through the sealing member inside the valve. In addition, it is possible to prevent leakage and evaporation of the fluid from the heat generated during the PCR reaction.
  • the first valve 300 may include a mixing chamber 315 including a nucleic acid binding material and a magnetic stirring bar 319 therein.
  • the first valve 300 includes a mixing chamber 315 therein to provide a space in which a sample and each reagent can be introduced and mixed.
  • the stirring process in the mixing chamber 315 is a process of mixing the sample and the reagent as part of the pre-processing process of removing and separating interfering substances from the sample before extracting the nucleic acid. After the sample containing the substance to be diagnosed and the reagent are introduced, they are stored in the mixing chamber 315 in the first valve 300 , respectively. A mixing process is performed, and thereafter, a substance to be diagnosed can be separated by a reagent.
  • the magnetic stir bar 319 is located inside the mixing chamber 315 of the first valve 300 and can serve to evenly mix the reagent, thereby accelerating the mixing of the sample and the reagent. can Accordingly, the mixing speed and efficiency are increased, and the total time required for the pretreatment process can be shortened. In this case, the rotation speed may be adjusted according to the sample to be pretreated.
  • the nucleic acid-binding material may serve to release nucleic acids from agglomeration, and may be silica beads.
  • the nucleic acid-binding material may be a magnetic bead having a magnetism, and the surface may be coated with silica or a carboxyl group.
  • the bead should be provided with an amino acid series that will halve the toxic substances of the metal by having a magnetic core in the core, and enveloping the core in the outermost layer of the bead, which will bind DNA or cells to be extracted on the outermost surface of the bead. Not limited.
  • a magnet part 370 capable of vertical movement, horizontal movement, or rotation movement may be provided under the first valve.
  • a magnetic force may be applied to the first valve 300 to separate the interfering material from the nucleic acid attached to the surface of the magnetic silica bead.
  • the magnet unit is disposed adjacent to the lower portion of the first valve 300 and serves to attract the magnetic beads.
  • the magnet part can be vertically moved, horizontally moved, or rotated under the first valve 300 , and the shape thereof is not limited.
  • the magnet part may include a support part for supporting the magnet.
  • the support part may be connected to the pretreatment chip to support the magnet part.
  • the magnet part may include an elastic member for shock mitigation between the magnet and the support part, and the elastic member may be preferably a spring.
  • a waste collecting part (not shown) that collects the separated interfering material after mixing by the rotational motion of the magnetic stir bar 319 is the lower portion of the first plate 210 or It may be provided under the mixing chamber 315 and may be in the form of a film. There is no need to manually remove the separated interfering material after the stirring process, and it can be recovered through the waste collecting part, thereby reducing the time required for the pretreatment process and easily separating the nucleic acid material to be diagnosed There are advantages to doing.
  • a bubble removing unit (not shown) may be further included in the first valve 300 .
  • the bubble removing unit may be disposed on the inner upper surface of the mixing chamber 315 and protrude toward the first plate 210 .
  • the bubbles contained in the fluid are pushed to the surroundings by the bubble removal unit due to buoyancy and are arranged in the surrounding space.
  • the bubble removal unit is exemplary, and the bubble removal unit may be used for various purposes according to an embodiment to which the present invention is applied.
  • the bubble removal unit may be used to remove bubbles included in the fluid from the flow of the fluid while the fluid passes through the reaction region.
  • the second valve 400 may include a distribution chamber 415 therein and may be controlled so that the fluid containing the nucleic acid is evenly distributed in the plurality of third channels 239 .
  • the fluid is a nucleic acid, for example, double-stranded DNA, an oligonucleotide primer having a sequence complementary to a specific nucleotide sequence to be amplified, a DNA polymerase, deoxyribonucleotide triphosphates (dNTP), and a PCR reaction buffer (PCR). reaction buffer) and the like.
  • the third channel 239 corresponds to the plurality of PCR reaction channels 530 and has one end connected to the second valve 400 to transfer nucleic acids to the plurality of PCR reaction channels 530 .
  • the distribution chamber serves to guide the introduced fluid. Due to the second valve 400, there is an advantage that precise nucleic acid distribution to each of the PCR reaction channels 530 is possible.
  • the second valve is provided with the number of inlet portions corresponding to the second channel and the number of outlet portions corresponding to the third channel on the side, and the second valve by turning movement The opening and closing of the valve can be adjusted.

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  • Health & Medical Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Biochemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • Molecular Biology (AREA)
  • Clinical Laboratory Science (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Apparatus Associated With Microorganisms And Enzymes (AREA)

Abstract

La présente invention concerne une micropuce et son procédé de scellement, la micropuce comprenant : une entrée par laquelle un fluide est introduit; un ou plusieurs canaux de réaction dans lesquels une réaction prédéfinie est mise en oeuvre par rapport au fluide introduit; et une sortie par laquelle le fluide s'écoule hors du ou des canaux de réaction, l'entrée ou la sortie comprenant un élément de scellement.
PCT/KR2021/015738 2020-11-05 2021-11-02 Micropuce et son procédé de scellement WO2022098064A1 (fr)

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KR102233058B1 (ko) * 2020-11-05 2021-03-29 주식회사 미코바이오메드 마이크로 칩 및 이의 실링 방법

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JP2004513211A (ja) * 2000-10-31 2004-04-30 ポレックス，コーポレーション 超吸収性材料を含む自己密封式媒体
KR20170053288A (ko) * 2015-11-06 2017-05-16 연세대학교 산학협력단 마이크로칩과 이의 제조방법, 이를 이용한 효소 검출 시스템과 방법
KR101811026B1 (ko) * 2017-01-23 2017-12-20 한국과학기술원 유전자 판별칩
KR20180020408A (ko) * 2016-08-18 2018-02-28 나노바이오시스 주식회사 미세유체 칩의 입출구 구조 및 그의 밀봉 방법
KR20200014639A (ko) * 2018-08-01 2020-02-11 주식회사 미코바이오메드 복수의 열 블록을 구비한 핵산 증폭 장치
KR102233058B1 (ko) * 2020-11-05 2021-03-29 주식회사 미코바이오메드 마이크로 칩 및 이의 실링 방법

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KR20160058302A (ko) 2014-11-14 2016-05-25 나노바이오시스 주식회사 미세유체 칩의 실링 장치 및 그 동작 방법

Patent Citations (6)

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Publication number Priority date Publication date Assignee Title
JP2004513211A (ja) * 2000-10-31 2004-04-30 ポレックス，コーポレーション 超吸収性材料を含む自己密封式媒体
KR20170053288A (ko) * 2015-11-06 2017-05-16 연세대학교 산학협력단 마이크로칩과 이의 제조방법, 이를 이용한 효소 검출 시스템과 방법
KR20180020408A (ko) * 2016-08-18 2018-02-28 나노바이오시스 주식회사 미세유체 칩의 입출구 구조 및 그의 밀봉 방법
KR101811026B1 (ko) * 2017-01-23 2017-12-20 한국과학기술원 유전자 판별칩
KR20200014639A (ko) * 2018-08-01 2020-02-11 주식회사 미코바이오메드 복수의 열 블록을 구비한 핵산 증폭 장치
KR102233058B1 (ko) * 2020-11-05 2021-03-29 주식회사 미코바이오메드 마이크로 칩 및 이의 실링 방법

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