WO2022152244A1 - 一种环介导等温扩增芯片 - Google Patents

一种环介导等温扩增芯片 Download PDF

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Publication number
WO2022152244A1
WO2022152244A1 PCT/CN2022/072022 CN2022072022W WO2022152244A1 WO 2022152244 A1 WO2022152244 A1 WO 2022152244A1 CN 2022072022 W CN2022072022 W CN 2022072022W WO 2022152244 A1 WO2022152244 A1 WO 2022152244A1
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WIPO (PCT)
Prior art keywords
piston
chamber
loop
microchannel
isothermal amplification
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PCT/CN2022/072022
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English (en)
French (fr)
Inventor
颜菁
邹长华
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江苏汇先医药技术有限公司
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Publication of WO2022152244A1 publication Critical patent/WO2022152244A1/zh

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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M1/00—Apparatus for enzymology or microbiology
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M1/00—Apparatus for enzymology or microbiology
    • C12M1/04—Apparatus for enzymology or microbiology with gas introduction means
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M1/00—Apparatus for enzymology or microbiology
    • C12M1/34—Measuring or testing with condition measuring or sensing means, e.g. colony counters

Definitions

  • the utility model belongs to the technical field of nucleic acid detection, and relates to a loop-mediated isothermal amplification chip.
  • Microfluidics is a technology for precise control and manipulation of microscale fluids, which specifically integrates basic operating units such as sample preparation, reaction, separation, and detection in biological, chemical, and medical analysis processes into a microfluidic control unit of several square centimeters.
  • On-chip technology that automates the entire process of analysis. Due to its huge potential in biology, chemistry, medicine and other fields, it has developed into a new research field interdisciplinary in biology, chemistry, medicine, fluids, electronics, materials, machinery and other disciplines.
  • LAMP Loop-mediated isothermal amplification
  • LAMP technology provides conditions for in vitro amplification of nucleic acid fragments to exponentially amplify them in large quantities and adds fluorescent dyes or fluorescent markers during the nucleic acid amplification process, and uses an optical device to detect the intensity of the fluorescent signal. Analysis of the signal is the process by which nucleic acid amplification results are derived.
  • the current LAMP detector can integrate nucleic acid amplification and detection.
  • the reaction chamber usually the amplification reaction chamber
  • the reaction chip can be heated, Lighting, detection, etc. Therefore, there is a need for a loop-mediated isothermal amplification chip that can be adapted to a LAMP detector.
  • the purpose of the present utility model is to provide a loop-mediated isothermal amplification chip, which can realize fully automatic and rapid nucleic acid detection by the LAMP method in a fully sealed state.
  • a loop-mediated isothermal amplification chip includes:
  • a body which has a plurality of chambers and a plurality of microchannels communicating with the chambers, and the chambers include a sample loading chamber, a buffer chamber, a piston chamber and one or more reaction chambers;
  • control valve for connecting or blocking the sample adding chamber and the buffer chamber, connecting or blocking the sample adding chamber and the reaction chamber, and connecting the buffer chamber and the reaction chamber or blocking, at least part of the control valve is movably provided in the body, the control valve has a communication channel that can selectively communicate with at least part of the microchannel;
  • a piston for driving liquid to circulate between the chambers the piston is movably arranged in the piston chamber, and the piston chamber is divided by the piston into a first piston chamber portion with variable volume and a a second piston cavity, the first piston cavity communicates with the sample adding cavity, and the second piston cavity communicates with the buffer cavity;
  • the control valve has a first working position and a second working position. In the first working position, the sample adding chamber and the buffer chamber communicate with each other, and the sample adding chamber and the buffer chamber are all connected to each other. The reaction chamber is disconnected; in the second working position, the sample adding chamber and the reaction chamber are communicated with each other, and the buffer chamber is communicated with the reaction chamber.
  • the chamber, the microchannel and the communication channel are enclosed within the body.
  • the body is provided with a sample addition port communicating with the sample addition chamber and a cover plate capable of closing the sample addition port.
  • cover plate has elasticity, one end of the cover plate is fixed on the surface of the main body, and the other end of the cover plate is covered on the sample injection port.
  • the thickness of the weak portion is reduced and is thinner than both ends of the cover plate.
  • the sample loading chamber and the buffer chamber are located above the control valve, the reaction chamber is located below the control valve, and the piston is located on the side of the buffer chamber.
  • control valve is movably disposed through the body along the x-axis, the control valve having a first drive engagement end external to the body.
  • the piston is disposed in the piston cavity movably along the x-axis, and the piston is disposed on a piston rod extending along the x-axis and movable along the x-axis, the piston rod having an extension to a second drive engagement end outside the body.
  • the centerline of the piston rod and the centerline of the control valve are parallel to each other and do not coincide.
  • control valve and the piston rod are staggered to facilitate connection with the driving mechanism of the LAMP detector.
  • the plurality of microchannels include a first microchannel communicated with the sample addition chamber, a second microchannel communicated with the buffer chamber, and a third microchannel communicated with the reaction chamber
  • a microchannel and a fourth microchannel each of the reaction chambers is communicated with one of the third microchannels and one of the fourth microchannels, respectively, and the communication channel includes a channel for connecting the first microchannel and the fourth microchannel.
  • a first communication channel for connecting the second microchannel, a second communication channel for connecting the first microchannel and the third microchannel, and a second communication channel for connecting the second microchannel and the fourth microchannel The third communication channel communicated with the channel.
  • the first communication channel has an inlet and an outlet located on the upper surface of the control valve
  • the second communication channel and the third communication channel respectively have an inlet located on the upper surface of the control valve and an inlet located on the upper surface of the control valve. Outlet on the lower surface of the control valve.
  • the first piston cavity part and the sample adding cavity communicate with each other through a first gas flow channel
  • the second piston cavity part and the buffer cavity communicate with each other through a second gas flow channel
  • the number of the reaction chambers is multiple, the control valve has a plurality of the second working positions, and only one of the reaction chambers and the second working position is in each of the second working positions.
  • the sample adding chamber and the buffer chamber communicate with each other.
  • a loop-mediated isothermal amplification chip includes:
  • a body which has a reagent zone and a reaction zone;
  • a switch piston is located between the reagent zone and the reaction zone, the switch piston is movably arranged in the body, and a switch capable of connecting the reagent zone and the reaction zone is provided in the switch piston the first microchannel;
  • the switch piston has a first working state and a second working position. When in the first working position, the reagent zone and the reaction zone are communicated through the first microchannel; when in the second working position , the reagent zone and the reaction zone are blocked by the switch piston.
  • the loop-mediated isothermal amplification chip further includes a drive piston for mixing the fluid in the reagent zone or flowing the fluid in the reagent zone into the reaction zone.
  • the body is provided with a quantitative cavity
  • the driving piston is movably inserted into the quantitative cavity
  • the switching piston is provided with a second microchannel that can communicate with the quantitative cavity .
  • the reaction zone includes multiple LAMP reaction zones and at least one internal standard reaction zone, the number of the first microchannels is multiple, and each reaction zone corresponds to one of the first microchannels respectively.
  • a plurality of the reaction zones and the internal standard reaction zones are arranged at intervals along the x-axis direction, and the switch piston is movably inserted in the body along the x-axis.
  • the switching piston has a plurality of first working positions, and when the switching piston is in any first working position, only one of the reaction zones communicates with the reagent zone through the corresponding first microchannel.
  • the dosing chamber communicates with the reagent zone through a second microchannel.
  • a plurality of the first microchannels have a junction, and the second microchannels communicate with the junction.
  • the dosing chamber is isolated from the reaction zone when the switching piston is in the second operating position.
  • the switch piston is provided with an inlet corresponding to the reagent zone, an outlet corresponding to the reaction zone and a second inlet corresponding to the quantitative chamber.
  • the second inlet is an elongated hole extending along the moving direction of the switch piston.
  • the body is further provided with a vent in communication with the reagent zone.
  • the loop-mediated isothermal amplification chip further includes a sealing membrane for closing the vent.
  • the utility model adopts the above scheme, and has the following advantages compared with the prior art:
  • the loop-mediated isothermal amplification chip of the utility model can realize nucleic acid detection by the LAMP method, and the detection process can be carried out in a fully sealed state, which reduces the external interference to the detection environment, and also avoids toxic or harmful substances in the sample.
  • the aerosol method escapes to the outside, which is convenient and safe to use, and is conducive to the automation of nucleic acid detection using the LAMP method.
  • Fig. 1 is the outline schematic diagram of a kind of loop-mediated isothermal amplification chip according to embodiment 1;
  • FIG. 2 is a front view of a loop-mediated isothermal amplification chip according to Embodiment 1;
  • FIG. 3 and 4 are perspective views of a loop-mediated isothermal amplification chip according to Embodiment 1, respectively;
  • Fig. 5 is the schematic diagram of the driving piston of embodiment 1;
  • FIG. 6 is a schematic view of the switching piston of Example 1.
  • FIG. 7 is a schematic perspective view of a loop-mediated isothermal amplification chip according to Embodiment 2 of the present invention from one viewing angle;
  • FIG. 8 is a schematic perspective view of the loop-mediated isothermal amplification chip shown in FIG. 7 from another perspective;
  • FIG. 9 is a front view of the loop-mediated isothermal amplification chip shown in FIG. 7;
  • Figure 10 is a top view of the loop-mediated isothermal amplification chip shown in Figure 7;
  • Figure 11a, Figure 11b and Figure 11c show a top view, a front view and a bottom view of the control valve according to Embodiment 2 of the present invention
  • FIG. 12 is a schematic diagram of the communication between the sample loading chamber and the buffer chamber according to Embodiment 2 of the present utility model, wherein the control valve is in the first working position;
  • FIG. 13 is a schematic diagram of the communication of the sample addition chamber, the buffer chamber and the first reaction chamber in Embodiment 2 of the present utility model, wherein the control valve is in the first second working position;
  • Example 14 is a schematic diagram of the communication of the sample addition chamber, the buffer chamber and the second reaction chamber in Example 2 of the present utility model, wherein the control valve is in the second second working position;
  • FIG. 15 is a schematic diagram of the communication of the sample addition chamber, the buffer chamber and the third reaction chamber in Embodiment 2 of the present utility model, wherein the control valve is in the third second working position;
  • Fig. 16 is a schematic diagram of the communication between the sample addition chamber, the buffer chamber and the fourth reaction chamber according to the second embodiment of the present invention, wherein the control valve is in the fourth second working position.
  • 100 body; 11, sample addition chamber; 110, first microchannel; 12, buffer chamber; 120, second microchannel; 13a, first piston chamber; 13b, second piston chamber; 131, first airflow channel; 132, second airflow channel; 14, reaction chamber; 141, third microchannel; 142, fourth microchannel;
  • control valve 210, first drive engagement end; 211, first communication passage; 212, second communication passage; 213, third communication passage;
  • a loop-mediated isothermal amplification chip in this embodiment includes a body 1 and a switch piston 5 .
  • the body 1 has a plurality of chambers, and specifically includes a reagent area 2 and a reaction area 6 .
  • the reagent area 2 is arranged in the upper part of the main body 1 and is used for pre-filling reagents; specifically, it is pre-filled with neutralizing liquid.
  • the reaction zone 6 is arranged in the lower part of the main body 1 .
  • the switch piston 5 is located between the reagent zone 2 and the reaction zone 6 , and the switch piston 5 is movably arranged in the body 1 .
  • the reagent zone 2 and the reaction zone 6 are chambers opened in the body 1 ; the switch piston 5 is inserted into the body 1 so as to be horizontally movable along the x-axis, and the reagent zone 2 is above the switch piston 5 , the reaction zone 6 is below the switch piston 5 .
  • the switch piston 5 is provided with a first microchannel 51 that can communicate the reagent zone 2 and the reaction zone 6 .
  • the switch piston 5 has a first working position and a second working position. In the first working position, the reagent zone 2 and the reaction zone 6 are communicated through the first microchannel 51; in the second working position, the reagent zone 2 and the reaction zone 6 are connected Isolated by switch piston 5.
  • the loop-mediated isothermal amplification chip further includes a driving piston 4 for mixing the fluid in the reagent zone 2 or allowing the fluid in the reagent zone 2 to flow into the reaction zone 6 .
  • the main body 1 is provided with a quantitative cavity 3
  • the driving piston 4 is movably inserted into the quantitative cavity 3
  • the switch piston 5 is provided with a second microchannel 52 that can communicate with the quantitative cavity 3 .
  • the quantitative chamber 3 is located on the side of the reagent area 2 and above the switch piston 5 .
  • the switching piston 5 also has at least one position that enables the reagent zone 2 to communicate with the dosing chamber 3 .
  • a rubber plug 41 is provided on the lower part of the driving piston 4 .
  • the reaction zone 6 includes a plurality of LAMP reaction zones 61 and at least one internal standard reaction zone 62 , the number of the first microchannels 51 is multiple, and each reaction zone 6 corresponds to one first microchannel 51 respectively.
  • the switching piston 5 has a plurality of first working positions. When the switching piston 5 is in any first working position, only one of the reaction zones 6 communicates with the reagent zone 2 through the corresponding first microchannel 51 . When the switch piston 5 is in the first working position, the quantitative chamber 3 communicates with the reagent zone 2 through the second microchannel 52 .
  • the plurality of first microchannels 51 have intersections, and the second microchannels 52 are connected to the intersections of the first microchannels 51.
  • the upper and lower surfaces of the switch piston 5 are respectively provided with a plurality of functional holes.
  • Part of the functional holes on the upper surface are used as the inlet 501 of the corresponding reagent zone 2, that is, the inlet 501 of the first microchannel 51; the functional holes on the lower surface are used as the outlet of the corresponding reaction zone 6, that is, the outlet of the first microchannel 51 .
  • a functional hole on the upper surface is used as the first communication port 502 corresponding to the quantitative chamber 3, and another functional hole on the upper surface is used as the second communication port 503 corresponding to the reagent area 2.
  • the second microchannel 52 connects the first The communication port 502 communicates with the second communication port 503 .
  • the first communication port 502 is a long hole extending along the sliding direction of the switch piston 5 .
  • the second communication ports 503 and the inlets 501 of the plurality of first microchannels 51 are arranged at intervals along the moving direction of the switching piston 5 .
  • the main body 1 is also provided with a vent 21 communicating with the reagent area 2 .
  • the loop-mediated isothermal amplification chip also includes a sealing membrane for closing the vent.
  • the amplification reaction of LAMP reagent is realized by constant temperature heating, and then detected by the optical module to realize the amplification and detection of nucleic acid.
  • FIG. 1 to FIG. 10 show a loop-mediated isothermal amplification chip of this embodiment, which is referred to as a LAMP chip for short.
  • the LAMP chip can be loaded into a LAMP detector specially configured for it to carry out constant temperature amplification and detection, so as to realize the qualitative and quantitative detection of nucleic acid.
  • the loop-mediated isothermal amplification chip mainly includes a body 100 , a control valve 200 and a piston 300 .
  • the body 100 has a plurality of chambers and a plurality of microchannels, each of which is communicated with one or two chambers respectively, and the plurality of chambers includes a sample loading chamber 11, a buffer chamber 12, a piston chamber and a plurality of reaction chambers 14.
  • the control valve 200 has a communication channel that can selectively communicate with at least part of the microchannels, so that it can be used to communicate or block the sample adding chamber 11 and the buffer chamber 12, and connect or block the sample adding chamber 11 and the reaction chamber 14 , and connect or block the buffer chamber 12 and the reaction chamber 14 .
  • the piston 300 is used to drive the liquid to circulate between the chambers, the piston 300 is movably arranged in the above-mentioned piston chamber, and the piston chamber is divided by the piston 300 into a first piston chamber portion 13a and a second piston chamber portion 13b with variable volumes , the first piston cavity 13a communicates with the sample adding cavity 11 , and the second piston cavity 13b communicates with the buffer cavity 12 .
  • the control valve 200 has a first working position and a second working position.
  • the sample adding chamber 11 and the buffer chamber 12 communicate with each other, and the sample adding chamber 11 and the buffer chamber 12 are not connected with the reaction chamber 14;
  • the sample adding chamber 11 and the reaction chamber 14 communicate with each other, and the buffer chamber 12 communicates with the reaction chamber 14 .
  • the control valve 200 is placed in the first working position to mix the sample liquid evenly; when the sample needs to be distributed into the reaction chamber 14, the control valve 200 is placed in the second working position.
  • the working position is convenient for distributing the sample reaction solution into the reaction chamber 14 .
  • the body 100 is integrally formed of plastic, such as injection molding, and forms a plurality of cavities and microchannels inside.
  • the above-mentioned chambers, microchannels and communication channels are all enclosed in the main body 100, and the liquid and gas circulation spaces inside the chip are completely isolated from the atmosphere except during sample addition.
  • the body 100 is provided with a sample addition port communicating with the sample addition cavity 11 and a cover plate 400 capable of closing the sample addition port.
  • the cover plate 400 has elasticity, one end of the cover plate 400 is fixed on the surface of the main body 100 , and the other end of the cover plate 400 covers the sample introduction port. Specifically, as shown in FIG. 1 , FIG. 2 and FIG.
  • the middle part of the cover plate 400 has a weak part 401 , and the thickness of the weak part 401 is reduced and thinner than the left and right ends of the cover plate 400 .
  • the cover plate 400 can be elastically deformed at the weak portion 401 to expose the sample injection port; after the sample addition is completed, the cover plate 400 can be reset and pressed against the main body 100 after the external force disappears. on the front surface and seal the injection port.
  • the body 100 is entirely made of transparent material, or at least the front part and the lower part corresponding to the reaction chamber are made of transparent material.
  • the sample addition chamber 11 and the buffer chamber 12 are located above (preferably directly above) the control valve 200, the reaction chamber 14 is located below (preferably directly below) the control valve 200, and the piston 300 is located beside the buffer chamber 12 (specifically for the rear).
  • the control valve 200 is movably inserted into the body 100 along the x-axis, where the x-axis extends horizontally along the left-right direction.
  • the control valve 200 has a first driving engagement end 210 located outside the body 100 so as to be connected with the driving mechanism (eg, mechanical arm, etc.) of the LAMP detector, so as to be driven and controlled to move precisely.
  • the piston 300 is arranged in the piston cavity so as to be movable along the x-axis, and the piston 300 is fixedly arranged on a piston rod 31 extending along the x-axis and movable along the x-axis.
  • the piston rod 31 has a second driving engagement end 310 extending to the outside of the body 100 so as to be connected with the driving mechanism (eg, mechanical arm, etc.) of the LAMP detector, so as to be driven and controlled to move accurately.
  • the center line of the piston rod 31 and the center line of the control valve 200 are parallel to each other and do not coincide with each other. That is, when viewed from a plan view, the control valve 200 and the piston rod 31 are staggered.
  • first driving engagement end portion 210 and the second driving engagement end portion 310 are independently located on the left or right side of the body 100 , that is, the movement directions of the control valve 200 and the piston 300 are parallel, so as to facilitate the LAMP detection
  • the first drive engagement end 210 and the second drive engagement end are respectively provided with engagement slots for connecting with the drive mechanism of the LAMP detector.
  • the number of reaction chambers 14 is multiple, and the plurality of reaction chambers 14 are arranged at intervals along the x-axis.
  • the control valve 200 has a plurality of second working positions, and in each second working position, only one of the reaction chambers 14 and the sample adding chamber 11 and the buffer chamber 12 communicate with each other. Specifically, as shown in FIGS. 7 to 10 , in this embodiment, four reaction chambers 14 are provided, and the control valve 200 has four second working positions.
  • the plurality of microchannels on the main body 100 include a first microchannel 110 communicated with the sample loading chamber 11 , a second microchannel 120 communicated with the buffer chamber 12 , and a plurality of microchannels.
  • the plurality of third microchannels 141 and the plurality of fourth microchannels 142 are connected to each of the reaction chambers 14 .
  • Each reaction chamber 14 is communicated with a third microchannel 141 and a fourth microchannel 142 respectively.
  • the plurality of communication channels on the control valve 200 include a first communication channel 211 for connecting the first microchannel 110 and the second microchannel 120, a first communication channel 211 for connecting the first microchannel 110 and the second microchannel 120.
  • a plurality of second communication channels 212 communicated with the third microchannel 141 and a plurality of third communication channels 213 for connecting the second microchannel 120 and the fourth microchannel 142 with each other.
  • the first communication passage 211 is generally opened at the upper portion of the control valve 200 , so as to have an inlet and an outlet opened on the upper surface of the control valve 200 .
  • the second communication channel 212 corresponds to the third microchannel 141 one-to-one.
  • the second communication channel 212 penetrates the control valve 200 in the up-down direction, so as to have an inlet opened on the upper surface of the control valve 200 and an inlet opened on the lower surface of the control valve 200 Export.
  • the third communication channel 213 corresponds to the fourth microchannel 142 one-to-one, and the third communication channel penetrates the control valve 200 in the up-down direction, so as to have an inlet opened on the lower surface of the control valve 200 and an outlet opened on the upper surface of the control valve 200 .
  • the first piston cavity 13 a and the sample adding cavity 11 communicate with each other through the first airflow channel 131
  • the second piston cavity 13 b and the buffer cavity 12 communicate through the second airflow channel 132 .
  • the first airflow channel 131 and the second airflow channel 132 are opened on the upper part of the main body 100 .
  • the microchannels and air flow channels on the body 100 can be arranged inside the body 100 or on the surface of the body 100 , if they are arranged on the surface of the body 100 , they are sealed by a sealing film or a sealing plate.
  • loop-mediated isothermal amplification chip The use process of the loop-mediated isothermal amplification chip will be described in detail below.
  • the control valve 200 is moved to its first working position.
  • the first microchannel 110 , the first communication channel 211 and the second microchannel 120 are connected in sequence to connect the sample loading chamber 11 and the buffer chamber. 12 is connected, the sample loading chamber 11 is communicated with the first piston cavity 13a through the first gas flow channel 131, and the second piston cavity 13b is communicated with the buffer cavity 12 through the second gas flow channel 132, thereby forming a fluid circuit for gas and liquid circulation ;
  • the piston 300 By moving the piston 300 back and forth, the sample liquid is urged to flow between the sample adding chamber 11 and the buffer chamber 12, and the liquid is mixed evenly.
  • the second communication channel 212 and the first microchannel 110 are staggered from each other, and the third communication channel 213 and the second microchannel 120 are staggered from each other, that is, the reaction chamber 14 is blocked by the control valve 200 and does not communicate with the sample addition chamber 11 . communicates with the buffer chamber 12 .
  • the sample solution is distributed to each reaction chamber 14 in sequence. As shown in FIG. 7 , first move the control valve 200 to the left for a certain distance to the first and second working position. At this time, the inlet of the third microchannel 141 of the first reaction chamber 14 communicates with its corresponding second The outlet of the channel 212 is aligned and connected, the outlet of the fourth microchannel 142 of the first reaction chamber 14 is aligned and connected to the inlet of the corresponding third communication channel 213, and the outlet of the third communication channel 213 is aligned with the second microchannel 120 is in butt communication, the inlet of the second communication channel 212 is in butt communication with the first microchannel 110, the sample loading chamber 11 communicates with the first piston cavity 13a through the first airflow channel 131, and the second piston cavity 13b passes through the second airflow
  • the channel 132 communicates with the buffer chamber 12 to form a fluid circuit for gas and liquid circulation; by moving the piston 300 back and forth, the sample liquid is urged to flow into the first reaction chamber 14 from
  • the control valve 200 is moved to the left for a certain distance to the second second working position.
  • the inlet of the third microchannel 141 of the second reaction chamber 14 communicates with the corresponding second working position.
  • the outlet of the channel 212 is aligned and connected
  • the outlet of the fourth microchannel 142 of the second reaction chamber 14 is aligned and connected to the inlet of the corresponding third communication channel 213, and the outlet of the third communication channel 213 is aligned with the second microchannel 120 is in butt communication
  • the inlet of the second communication channel 212 is in butt communication with the first microchannel 110
  • the sample loading chamber 11 communicates with the first piston cavity 13a through the first airflow channel 131
  • the second piston cavity 13b passes through the second airflow
  • the channel 132 communicates with the buffer chamber 12 to form a fluid circuit for gas and liquid circulation; by moving the piston 300 back and forth, the sample liquid is urged to flow into the second reaction chamber 14 from the upper sample loading chamber 11 or the buffer chamber 12 . In this state,
  • the control valve 200 is moved to the left for a certain distance to the third second working position.
  • the inlet of the third microchannel 141 of the third reaction chamber 14 communicates with the corresponding second working position.
  • the outlet of the channel 212 is aligned and connected
  • the outlet of the fourth microchannel 142 of the third reaction chamber 14 is aligned and connected to the inlet of the corresponding third communication channel 213, and the communication mode of the sample addition chamber 11 and the buffer chamber 12 is the same as the previous one.
  • the description is similar, thereby forming a fluid circuit including the third reaction chamber 14 ; by moving the piston 300 back and forth, the sample liquid flows into the third reaction chamber 14 .
  • the control valve 200 is moved to the left for a certain distance to the fourth second working position.
  • the inlet of the third microchannel 141 of the fourth reaction chamber 14 is in communication with the corresponding second working position.
  • the outlet of the channel 212 is aligned and connected, the outlet of the fourth microchannel 142 of the fourth reaction chamber 14 is aligned and connected to the inlet of the corresponding third communication channel 213, and the communication mode of the sample loading chamber 11 and the buffer chamber 12 is the same as the previous one.
  • the description is similar, thereby forming a fluid circuit including the fourth reaction chamber 14 ; by moving the piston 300 back and forth, the sample liquid flows into the fourth reaction chamber 14 .
  • each reaction chamber 14 is heated at a constant temperature by the heating module of the LAMP detector to realize constant temperature amplification, and the optical module of the LAMP detector is used to perform fluorescence excitation on each reaction chamber 14 and collect fluorescence, according to the fluorescence color.
  • Qualitative analysis determination of negative/positive was performed, and quantitative analysis was performed according to the fluorescence value.

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Abstract

一种环介导等温扩增芯片，包括：本体（100），具有多个腔室及与多个微通道，腔室包括加样腔（11）、缓冲腔（12）、活塞腔及反应仓（14）；控制阀（200），用于将加样腔（11）和缓冲腔（12）连通或阻断、将加样腔（11）和反应仓（14）连通或阻断、及将缓冲腔（12）和反应仓（14）连通或阻断，控制阀（200）的至少部分活动地设于本体内；活塞（300），用于驱动液体在腔室之间流通，活塞（300）可移动地设置于活塞腔内，活塞腔被活塞分隔为容积可变的第一活塞腔部（13a）和第二活塞腔部（13b），第一活塞腔部（13a）和加样腔（11）连通，第二活塞腔部（13b）和缓冲腔（12）连通；控制阀200）在第一工作位置时，加样腔（11）和缓冲腔（12）相互连通；在第二工作位置时，加样腔（11）和反应仓（14）相互连通，缓冲腔（12）和反应仓（14）连通。所述芯片能够实现用LAMP方法在全密封状态下进行全自动及快速的核酸检测。

Description

一种环介导等温扩增芯片
相关申请的交叉引用
本申请要求2021年1月15日提交的申请号为CN202120109588X的中国专利申请的优先权，其全部内容通过引用的方式并入本发明中。
技术领域
本实用新型属于核酸检测技术领域，涉及一种环介导等温扩增芯片。
背景技术
微流控是一种精确控制和操控微尺度流体的技术，其具体为将生物、化学、医学分析过程的样品制备、反应、分离、检测等基本操作单元集成到一块几平方厘米的微流控芯片上，自动完成分析全过程的技术。由于其在生物、化学、医学等领域的巨大潜力，已经发展成为一个生物、化学、医学、流体、电子、材料、机械等学科交叉的崭新研究领域。
LAMP(环介导等温扩增，Loop-mediated isothermal amplification)技术因其反应条件温和(反应温度较低)、反应时间短等优点广泛应用于生物诊断领域，如进行核酸扩增检测以诊断样本中是否存在病原体。LAMP技术是通过为核酸片段提供体外扩增的条件，使之成指数大量扩增并在核酸扩增过程中加入荧光染料或荧光标记物，采用光学装置检测出荧光信号的强弱，通过对荧光信号的分析得出核酸扩增结果的过程。目前的LAMP检测仪能够集核酸扩增检测于一体，当检测芯片(通常为微流控芯片)放入LAMP检测仪后，能够对检测芯片的反应仓(通常为扩增反应仓)进行加热、光照、检测等。因此，需要一种能够适配LAMP检测仪的环介导等温扩增芯片。
实用新型内容
本实用新型的目的是提供一种环介导等温扩增芯片，可以实现用LAMP方法在全密封状态下进行全自动及快速的核酸检测。
根据本发明的第一个方面，一种环介导等温扩增芯片，包括：
本体，其具有多个腔室及多个与所述腔室连通的微通道，所述腔室包括加样腔、缓冲腔、活塞腔及一或多个反应仓；
控制阀，其用于将所述加样腔和所述缓冲腔连通或阻断、将所述加样腔和所述反应仓连通或阻断、及将所述缓冲腔和所述反应仓连通或阻断，所述控制阀的至少部分活动地设于所述本体内，所述控制阀具有能够选择性地与至少部分的所述微通道连通的连通通道；
活塞，其用于驱动液体在所述腔室之间流通，所述活塞可移动地设置于所述活塞腔内，所述活塞腔被所述活塞分隔为容积可变的第一活塞腔部和第二活塞腔部，所述第一活塞腔部和所述加样腔连通，所述第二活塞腔部和所述缓冲腔连通；
所述控制阀具有第一工作位置和第二工作位置，在所述第一工作位置时，所述加样腔和所述缓冲腔相互连通，所述加样腔、所述缓冲腔均和所述反应仓不连通；在所述第二工作位置时，所述加样腔和所述反应仓相互连通，所述缓冲腔和所述反应仓连通。
在一些实施例中，所述腔室、所述微通道及所述连通通道被封闭在所述本体内。
在一些更优选的实施例中，所述本体上设有与所述加样腔连通的加样口及能够将所述加样口封闭的盖板。
进一步地，所述盖板具有弹性且其一端部固定在所述本体的表面上，所述盖板的另一端部覆在所述加样口上。
更进一步地，该薄弱部的厚度减小，比盖板的两端部薄。
在一些实施例中，所述加样腔和所述缓冲腔位于所述控制阀的上方，所述反应仓位于所述控制阀的下方，所述活塞位于所述缓冲腔的旁侧。
在一些实施例中，所述控制阀可沿x轴移动地穿设于所述本体中，所述控制阀具有位于所述本体外部的第一驱动接合端部。
在一些实施例中，所述活塞可沿x轴移动地设于所述活塞腔内，所述活塞设置于一沿x轴延伸且可沿x轴移动的活塞杆上，所述活塞杆具有延伸至所述本体外部的第二驱动接合端部。
优选地，活塞杆的中心线和控制阀的中心线相互平行且不相重合。
具体地，在俯视时，控制阀和活塞杆错开设置，以便于和LAMP检测仪的驱动机构连接。
在一些更优选的实施例中，所述多个微通道包括与所述加样腔连通的第一微通道、与所述缓冲腔连通的第二微通道、与所述反应仓连通的第三微通道及第四微通道，每个所述反应仓分别与一个所述第三微通道及一个所述第四微通道连通，所述连通通道包括用于将所述第一微通道和所述第二微通道连通的第一连通通道、用于将所述第一微通道和所述第三微通道连通的第二连通通道及用于将所述第二微通道及所述第四微通道道连通的第三连通通道。
进一步地，所述第一连通通道具有位于所述控制阀上表面的入口和出口，所述第二连通通道和所述第三连通通道分别具有位于所述控制阀上表面的入口及位于所述控制阀下表面的出口。
在一些实施例中，所述第一活塞腔部和所述加样腔通过第一气流通道连通，所述第二活塞腔部和所述缓冲腔通过第二气流通道连通。
在一些实施例中，所述反应仓的数量为多个，所述控制阀具有多个所述第二工作位置，在 每个所述第二工作位置时仅其中一个所述反应仓和所述加样腔及所述缓冲腔相互连通。
根据本发明的第二个方面，一种环介导等温扩增芯片，包括：
本体，其具有试剂区和反应区；
开关活塞，其位于所述试剂区和所述反应区之间，所述开关活塞活动地设于所述本体内，所述开关活塞中设有能够将所述试剂区和所述反应区连通的第一微通道；
所述开关活塞具有第一工作状态和第二工作位置，在所述第一工作位置时，所述试剂区和所述反应区通过所述第一微通道连通；在所述第二工作位置时，所述试剂区和所述反应区被所述开关活塞阻断。
在一些实施例中，所述环介导等温扩增芯片还包括用于使所述试剂区内的流体混匀或使所述试剂区内的流体流入所述反应区的驱动活塞。
在一些实施例中，所述本体上设有定量腔，所述驱动活塞可移动地插设于所述定量腔内，所述开关活塞上设置有能够和所述定量腔连通的第二微通道。
在一些实施例中，所述反应区包括多个LAMP反应区和至少一个内标反应区，所述第一微通道的数量为多个，每个反应区分别对应一个所述第一微通道。
在一些优选的实施例中，多个所述反应区和所述内标反应区沿x轴方向间隔排布，所述开关活塞能够沿x轴移动地插设于所述本体中。
在一些实施例中，所述开关活塞具有多个第一工作位置，当所述开关活塞在任一第一工作位置时，仅其中一个反应区通过相应的第一微通道和所述试剂区连通。
在一些实施例中，当所述开关活塞在第一工作位置时，所述定量腔通过第二微通道和所述试剂区相互连通。
在一些优选的实施例中，多个所述第一微通道具有交汇处，所述第二微通道与所述交汇处连通。
在一些实施例中，当所述开关活塞在第二工作位置时，所述定量腔与所述反应区相隔离。
在一些实施例中，所述开关活塞上设有对应所述试剂区的入口、对应所述反应区的出口及对应所述定量腔的第二入口。
在一些优选的实施例中，所述第二入口为沿所述开关活塞的移动方向延伸的长孔。
在一些实施例中，所述本体上还设有与所述试剂区连通的通气口。
在一些实施例中，所述环介导等温扩增芯片还包括用于封闭所述通气口的密封膜。
本实用新型采用以上方案，相比现有技术具有如下优点：
本实用新型的环介导等温扩增芯片可以实现用LAMP方法进行核酸检测，检测过程可以 在全密封状态下进行，减少了外部对检测环境的干扰，同时也避免了样本中有毒或有害物质以气溶胶方式逸散到外部，使用方便且安全，利于采用LAMP方法实现核酸检测的自动化。
附图说明
为了更清楚地说明本实用新型的技术方案，下面将对实施例描述中所需要使用的附图作简单地介绍，显而易见地，下面描述中的附图仅仅是本实用新型的一些实施例，对于本领域普通技术人员来讲，在不付出创造性劳动的前提下，还可以根据这些附图获得其他的附图。
图1是根据实施例1的一种环介导等温扩增芯片的外形示意图；
图2是根据实施例1的一种环介导等温扩增芯片的主视图；
图3、4分别是根据实施例1的一种环介导等温扩增芯片的透视图；
图5是实施例1的驱动活塞的示意图；
图6是实施例1的开关活塞的示意图。
图7是根据本实用新型实施例2的一种环介导等温扩增芯片在一个视角下的立体示意图；
图8是图7所示环介导等温扩增芯片在另一个视角下的立体示意图；
图9是图7所示环介导等温扩增芯片的主视图；
图10是图7所示环介导等温扩增芯片的俯视图；
图11a、图11b及图11c示出了本实用新型实施例2的控制阀的俯视图、主视图及仰视图；
图12为本实用新型实施例2的加样腔和缓冲腔的连通示意图，其中控制阀处于第一工作位置；
图13为本实用新型实施例2的加样腔、缓冲腔和第一个反应仓的连通示意图，其中控制阀处于第一个第二工作位置；
图14为本实用新型实施例2的加样腔、缓冲腔和第二个反应仓的连通示意图，其中控制阀处于第二个第二工作位置；
图15为本实用新型实施例2的加样腔、缓冲腔和第三个反应仓的连通示意图，其中控制阀处于第三个第二工作位置；
图16为本实用新型实施例2的加样腔、缓冲腔和第四个反应仓的连通示意图，其中控制阀处于第四个第二工作位置。
其中，
1、本体；2、试剂区；21、通气口；3、定量腔；4、驱动活塞；41、橡胶塞；5、开关活塞；501、入口；502、第一连通口；503、第二连通口；51、第一微通道；52、第二微通道；6、 反应区；61、LAMP反应区；62、内标反应区。
100、本体；11、加样腔；110、第一微通道；12、缓冲腔；120、第二微通道；13a、第一活塞腔部；13b、第二活塞腔部；131、第一气流通道；132、第二气流通道；14、反应仓；141、第三微通道；142、第四微通道；
200、控制阀；210、第一驱动接合端部；211、第一连通通道；212、第二连通通道；213、第三连通通道；
300、活塞；31、活塞杆；310、第二驱动接合端部；
400、盖板；401、薄弱部。
具体实施方式
下面结合附图对本实用新型的较佳实施例进行详细阐述，以使本实用新型的优点和特征能更易于被本领域的技术人员理解。在此需要说明的是，对于这些实施方式的说明用于帮助理解本实用新型，但并不构成对本实用新型的限定。
实施例1
参照图1至图6所示，本实施例的一种环介导等温扩增芯片(简称LAMP芯片)，包括：本体1和开关活塞5。本体1具有多个腔室，具体包括试剂区2、反应区6。试剂区2设于本体1的上部内，并用于预灌装试剂；具体预灌装有中和液。反应区6设于本体1的下部内。开关活塞5位于试剂区2和反应区6之间，开关活塞5可移动地设于本体1内。具体到本实施例中，试剂区2和反应区6为开设于本体1中的腔室；开关活塞5可沿x轴水平移动地插设于本体1中，试剂区2在开关活塞5的上方，反应区6在开关活塞5的下方。开关活塞5中设有能够将试剂区2和反应区6连通的第一微通道51。开关活塞5具有第一工作位置和第二工作位置，在第一工作位置时，试剂区2和反应区6通过第一微通道51连通；在第二工作位置时，试剂区2和反应区6由开关活塞5隔离。
该环介导等温扩增芯片还包括用于使试剂区2内的流体混匀或使试剂区2内的流体流入反应区6的驱动活塞4。本体1上设有定量腔3，驱动活塞4可移动地插设于定量腔3内，开关活塞5上设置有能够和定量腔3连通的第二微通道52。具体到本实施例中，定量腔3位于试剂区2的旁侧并位于开关活塞5的上方。开关活塞5还具有至少一个能够使试剂区2和定量腔3连通的位置。驱动活塞4的下部上设有橡胶塞41。
反应区6包括多个LAMP反应区61和至少一个内标反应区62，第一微通道51的数量为多个，每个反应区6分别对应一个第一微通道51。开关活塞5具有多个第一工作位置，当开关活塞5在任一第一工作位置时，仅其中一个反应区6通过相应的第一微通道51和试剂区2连通。当开关活塞5在第一工作位置时，定量腔3通过第二微通道52和试剂区2相互连通。具体到本实施例中，多个第一微通道51具有交汇处，第二微通道52连通第一微通道51的交汇处，当开关活塞5在任一个第一工作位置时，仅一个LAMP反应区61或内标反应区62通 过对应的第一微通道51连通至试剂区2，同时定量腔3也通过第二微通道52和试剂区2及该LAMP反应区61或内标反应区62连通，此时，向上移动驱动活塞4，可将试剂区2内的反应液抽出；再向下移动驱动活塞4，可将抽出的反应液推入处于连通状态的反应区6内。
如图6所示，开关活塞5的上下表面上分别设有多个功能孔。上表面上的部分功能孔用作对应试剂区2的入口501，即第一微通道51的入口501；下表面上的功能孔用作对应反应区6的出口，即第一微通道51的出口。上表面上的一个功能孔用作对应定量腔3的第一连通口502，上表面上还有另一个功能孔用作对应试剂区2的第二连通口503，第二微通道52将第一连通口502和第二连通口503连通。第一连通口502具体为沿开关活塞5的滑动方向延伸的长孔。第二连通口503和多个第一微通道51的入口501沿开关活塞5的移动方向间隔排列。
本体1上还设有与试剂区2连通的通气口21。该环介导等温扩增芯片还包括用于封闭通气口的密封膜。
上述LAMP芯片的操作步骤如下：
1.将样本充分裂解的保存液分别加入试剂区2(中和液预灌装区域)内。
2.左右移动开关活塞5，将试剂区2与定量腔3(混匀定量腔3)连通，通过上下移动驱动活塞4(混匀定量活塞)进行中和液和保存液的混匀反应，实现核酸提取。
3.左右移动开关活塞5将试剂区2、LAMP定量反应区6和内标反应区62的孔及定量腔3(混匀定量腔3)接通，通过上下移动驱动活塞4实现液体定量流入LAMP定量反应区6的孔内。
4.依次实现三个LAMP定量反应区6和一个内标反应区62的试剂定量。
5.通过移动开关活塞5将三个LAMP定量反应区6和一个内标反应区62关闭，与试剂区2、定量腔3之间隔开。
6.通过恒温加热实现LAMP试剂扩增反应，然后通过光学模块进行检测，实现对核酸的扩增检测。
实施例2
图1至图10示出了本实施例的一种环介导等温扩增芯片，简称LAMP芯片。该LAMP芯片能够装入为其专门配置的LAMP检测仪中进行恒温扩增和检测，实现对核酸的定性及定量检测。参照图1至图10所示，该环介导等温扩增芯片主要包括本体100、控制阀200和活塞300。本体100具有多个腔室及与多个微通道，每个微通道分别与一个或两个腔室连通，该多个腔室包括加样腔11、缓冲腔12、活塞腔及多个反应仓14。控制阀200具有能够选择性地与至少部分的微通道连通的连通通道，从而能够用于将加样腔11和缓冲腔12连通或阻断、将加样腔11和反应仓14连通或阻断、及将缓冲腔12和反应仓14连通或阻断。活塞300用于驱动液体在腔室之间流通，活塞300可移动地设置于上述的活塞腔内，活塞腔被活塞300分隔为容积可变的第一活塞腔部13a和第二活塞腔部13b，第一活塞腔部13a和加样腔11连 通，第二活塞腔部13b和缓冲腔12连通。该控制阀200具有第一工作位置和第二工作位置，在第一工作位置时，加样腔11和缓冲腔12相互连通，加样腔11、缓冲腔12均和反应仓14不连通；在第二工作位置时，加样腔11和反应仓14相互连通，缓冲腔12和反应仓14连通。具体来说，当加样完成及装入试剂后，使控制阀200处于第一工作位置，以使样本液混匀；当需要将样本分配到反应仓14中时，使控制阀200处于第二工作位置，以便于将样本反应液分配到反应仓14中。
本体100由塑料一体成型，如注塑成型，并在内部形成多个腔室和微通道。上述的各腔室、微通道及连通通道均被封闭在本体100内，除加样时，芯片内部的液体及气体流通空间和大气完全隔绝。本体100上设有与加样腔11连通的加样口及能够将加样口封闭的盖板400。盖板400具有弹性且其一端部固定在本体100的表面上，盖板400的另一端部覆在加样口上。具体如图1、图2和图4所示，该盖板400的中部具有薄弱部401，该薄弱部401的厚度减小，比盖板400的左右两端部薄。当向外拨盖板400的左端部时，盖板400可在该薄弱部401弹性变形，而将加样口露出；加样完成后，外力消失后盖板400即可复位压紧在本体100的前表面上而将加样口封住。本体100整体由透明材料制成，或至少对应反应仓的前侧部分及下侧部分由透明材料制成。
加样腔11和缓冲腔12位于控制阀200的上方(优选为正上方)，反应仓14位于控制阀200的下方((优选为正下方))，活塞300位于缓冲腔12的旁侧(具体为后侧)。控制阀200可沿x轴移动地穿设于本体100中，此处，x轴沿左右方向水平延伸。控制阀200具有位于本体100外部的第一驱动接合端部210，以便于和LAMP检测仪的驱动机构(如机械臂等)连接，从而能够被驱动而被控制精确移动。活塞300可沿x轴移动地设于活塞腔内，活塞300固定设置于一沿x轴延伸且可沿x轴移动的活塞杆31上。活塞杆31具有延伸至本体100外部的第二驱动接合端部310，以便于和LAMP检测仪的驱动机构(如机械臂等)连接，从而能够被驱动而被控制精确移动。活塞杆31的中心线和控制阀200的中心线相互平行且不相重合。也就是说，从俯视角度看，控制阀200和活塞杆31错开设置。还需要说明的是，第一驱动接合端部210和第二驱动接合端部310独立地位于本体100的左侧或右侧，即控制阀200和活塞300的移动方向平行，从而能够便于LAMP检测仪的驱动机构的布局和配置。第一驱动接合端部210和第二驱动接合端分别设有用于和LAMP检测仪的驱动机构连接的接合卡槽。
反应仓14的数量为多个，多个反应仓14沿x轴间隔排列。控制阀200具有多个第二工作位置，在每个第二工作位置时仅其中一个反应仓14和加样腔11及缓冲腔12相互连通。具体如图7至图10所示，本实施例中，设有四个反应仓14，控制阀200具有四个第二工作位置。
结合图4及图6至图10所示，本体100上的多个微通道包括与加样腔11连通的一个第一微通道110、与缓冲腔12连通的一个第二微通道120、与多个反应仓14连通的多个第三微通道141及多个第四微通道142。每个反应仓14分别与一个第三微通道141及一个第四微通道142连通。结合图3a至图3c所示，控制阀200上的多个连通通道包括用于将第一微通道 110和第二微通道120连通的第一连通通道211、用于将第一微通道110和第三微通道141连通的多个第二连通通道212及用于将第二微通道120及第四微通道142道连通的多个第三连通通道213。第一连通通道211大体开设于控制阀200的上部，从而具有开设于控制阀200上表面上的入口和出口。第二连通通道212与第三微通道141一一对应，第二连通通道212沿上下方向贯穿控制阀200，从而具有开设于控制阀200上表面上的入口和开设于控制阀200下表面上的出口。第三连通通道213与第四微通道142一一对应，第三连通道沿上下方向贯穿控制阀200，从而具有开设于控制阀200下表面上的入口和开设于控制阀200上表面上的出口。
结合图4及图6至图10所示，第一活塞腔部13a和加样腔11通过第一气流通道131连通，第二活塞腔部13b和缓冲腔12通过第二气流通道132连通。第一气流通道131和第二气流通道132开设于本体100的上部。本体100上的微通道和气流通道可设于本体100的内部或本体100的表面，若设于本体100的表面，则通过密封膜或密封板封住。
下面对该环介导等温扩增芯片的使用过程进行详细描述。
如图6所示，使控制阀200移动到其第一工作位置，此时，第一微通道110、第一连通通道211和第二微通道120依次对接连通从而将加样腔11和缓冲腔12连通，加样腔11通过第一气流通道131和第一活塞腔部13a连通，第二活塞腔部13b通过第二气流通道132和缓冲腔12连通，从而构成供气体和液体流通的流体回路；通过往复移动活塞300，促使样本液在加样腔11和缓冲腔12之间流动，促使液体混匀。在该状态下，第二连通通道212和第一微通道110相互错开，第三连通通道213和第二微通道120相互错开，即反应仓14被控制阀200阻断而不和加样腔11及缓冲腔12连通。
混匀完成后，依次将样本液分配到各反应仓14中。如图7所示，先使控制阀200向左移动一定距离至其第一个第二工作位置，此时，第一个反应仓14的第三微通道141的入口和与其对应的第二连通通道212的出口对齐相接，第一个反应仓14的第四微通道142的出口和与其对应的第三连通通道213的入口对齐相接，该第三连通通道213的出口和第二微通道120对接连通，该第二连通通道212的入口和第一微通道110对接连通，加样腔11通过第一气流通道131和第一活塞腔部13a连通，第二活塞腔部13b通过第二气流通道132和缓冲腔12连通，从而构成供气体和液体流通的流体回路；通过往复移动活塞300，促使样本液自上部的加样腔11或缓冲腔12流入第一个反应仓14。在该状态下，其他反应仓14被控制阀200阻断而不和加样腔11及缓冲腔12连通。
如图8所示，使控制阀200继续向左移动一定距离至其第二个第二工作位置，此时，第二个反应仓14的第三微通道141的入口和与其对应的第二连通通道212的出口对齐相接，第二个反应仓14的第四微通道142的出口和与其对应的第三连通通道213的入口对齐相接，该第三连通通道213的出口和第二微通道120对接连通，该第二连通通道212的入口和第一微通道110对接连通，加样腔11通过第一气流通道131和第一活塞腔部13a连通，第二活塞腔部13b通过第二气流通道132和缓冲腔12连通，从而构成供气体和液体流通的流体回路； 通过往复移动活塞300，促使样本液自上部的加样腔11或缓冲腔12流入第二个反应仓14。在该状态下，其他反应仓14被控制阀200阻断而不和加样腔11及缓冲腔12连通。
如图9所示，使控制阀200继续向左移动一定距离至其第三个第二工作位置，此时，第三个反应仓14的第三微通道141的入口和与其对应的第二连通通道212的出口对齐相接，第三个反应仓14的第四微通道142的出口和与其对应的第三连通通道213的入口对齐相接，加样腔11和缓冲腔12的连通方式与前文所述类似，从而构成包括第三个反应仓14在内的流体回路；通过往复移动活塞300，使样本液流入第三个反应仓14。
如图10所示，使控制阀200继续向左移动一定距离至其第四个第二工作位置，此时，第四个反应仓14的第三微通道141的入口和与其对应的第二连通通道212的出口对齐相接，第四个反应仓14的第四微通道142的出口和与其对应的第三连通通道213的入口对齐相接，加样腔11和缓冲腔12的连通方式与前文所述类似，从而构成包括第四个反应仓14在内的流体回路；通过往复移动活塞300，使样本液流入第四个反应仓14。
样本液分配完成后，通过LAMP检测仪的加热模块对各反应仓14进行恒温加热，实现恒温扩增，并通过LAMP检测仪的光学模块对各个反应仓14进行荧光激发并收集荧光，根据荧光颜色进行定性分析(判断阴性/阳性)，根据荧光值进行定量分析。
如本说明书和权利要求书中所示，术语“包括”与“包含”仅提示包括已明确标识的步骤和元素，而这些步骤和元素不构成一个排它性的罗列，方法或者设备也可能包含其他的步骤或元素。本文所使用的术语“和/或”包括一个或多个相关的所列项目的任意的组合。
需要说明的是，如无特殊说明，当某一特征被称为“固定”、“连接”在另一个特征，它可以直接固定、连接在另一个特征上，也可以间接地固定、连接在另一个特征上。此外，本实用新型中所使用的上、下、左、右等描述仅仅是相对于附图中本实用新型各组成部分的相互位置关系来说的。
进一步可以理解的是，本公开中“多个”是指两个或两个以上，其它量词与之类似。
上述实施例只为说明本发明的技术构思及特点，是一种优选的实施例，其目的在于熟悉此项技术的人士能够了解本发明的内容并据以实施，并不能以此限定本发明的保护范围。

Claims (21)

  1. 一种环介导等温扩增芯片，其特征在于，包括：
    本体，其具有多个腔室及多个与所述腔室连通的微通道，所述腔室包括加样腔、缓冲腔、活塞腔及一或多个反应仓；
    控制阀，其用于将所述加样腔和所述缓冲腔连通或阻断、将所述加样腔和所述反应仓连通或阻断、及将所述缓冲腔和所述反应仓连通或阻断，所述控制阀的至少部分活动地设于所述本体内，所述控制阀具有能够选择性地与至少部分的所述微通道连通的连通通道；
    活塞，其用于驱动液体在所述腔室之间流通，所述活塞可移动地设置于所述活塞腔内，所述活塞腔被所述活塞分隔为容积可变的第一活塞腔部和第二活塞腔部，所述第一活塞腔部和所述加样腔连通，所述第二活塞腔部和所述缓冲腔连通；
    所述控制阀具有第一工作位置和第二工作位置，在所述第一工作位置时，所述加样腔和所述缓冲腔相互连通，所述加样腔、所述缓冲腔均和所述反应仓不连通；在所述第二工作位置时，所述加样腔和所述反应仓相互连通，所述缓冲腔和所述反应仓连通。
  2. 根据权利要求1所述的环介导等温扩增芯片，其特征在于，所述腔室、所述微通道及所述连通通道被封闭在所述本体内。
  3. 根据权利要求2所述的环介导等温扩增芯片，其特征在于，所述本体上设有与所述加样腔连通的加样口及能够将所述加样口封闭的盖板。
  4. 根据权利要求3所述的环介导等温扩增芯片，其特征在于，所述盖板具有弹性且其一端部固定在所述本体的表面上，所述盖板的另一端部覆在所述加样口上。
  5. 根据权利要求1所述的环介导等温扩增芯片，其特征在于，所述加样腔和所述缓冲腔位于所述控制阀的上方，所述反应仓位于所述控制阀的下方，所述活塞位于所述缓冲腔的旁侧。
  6. 根据权利要求1或5所述的环介导等温扩增芯片，其特征在于，所述控制阀可沿x轴移动地穿设于所述本体中，所述控制阀具有位于所述本体外部的第一驱动接合端部；和/或，所述活塞可沿x轴移动地设于所述活塞腔内，所述活塞设置于一沿x轴延伸且可沿x轴移动的活塞杆上，所述活塞杆具有延伸至所述本体外部的第二驱动接合端部。
  7. 根据权利要求6所述的环介导等温扩增芯片，其特征在于，所述多个微通道包括与所述加样腔连通的第一微通道、与所述缓冲腔连通的第二微通道、与所述反应仓连通的第三微通道及第四微通道，每个所述反应仓分别与一个所述第三微通道及一个所述第四微通道连通，所述连通通道包括用于将所述第一微通道和所述第二微通道连通的第一连通通道、用于将所述第一微通道和所述第三微通道连通的第二连通通道及用于将所述第二微通道及所述第四微通道道连通的第三连通通道。
  8. 根据权利要求7所述的环介导等温扩增芯片，其特征在于，所述第一连通通道具有位于所述控制阀上表面的入口和出口，所述第二连通通道和所述第三连通通道分别具有位于所述控制阀上表面的入口及位于所述控制阀下表面的出口。
  9. 根据权利要求1所述的环介导等温扩增芯片，其特征在于，所述第一活塞腔部和所述加样腔通过第一气流通道连通，所述第二活塞腔部和所述缓冲腔通过第二气流通道连通。
  10. 根据权利要求1所述的环介导等温扩增芯片，其特征在于，所述反应仓的数量为多个，所述控制阀具有多个所述第二工作位置，在每个所述第二工作位置时仅其中一个所述反应仓和所述加样腔及所述缓冲腔相互连通。
  11. 一种环介导等温扩增芯片，其特征在于，包括：
    本体，其具有试剂区和反应区；
    开关活塞，其位于所述试剂区和所述反应区之间，所述开关活塞活地设于所述本体内，所述开关活塞中设有能够将所述试剂区和所述反应区连通的第一微通道；
    所述开关活塞具有第一工作位置和第二工作位置，在所述第一工作位置时，所述试剂区和所述反应区通过所述第一微通道连通；在所述第二工作位置时，所述试剂区和所述反应区被所述开关活塞阻断。
  12. 根据权利要求11所述的环介导等温扩增芯片，其特征在于，所述环介导等温扩增芯片还包括用于使所述试剂区内的流体混匀或使所述试剂区内的流体流入所述反应区的驱动活塞。
  13. 根据权利要求12所述的环介导等温扩增芯片，其特征在于，所述本体上设有定量腔，所述驱动活塞可移动地插设于所述定量腔内，所述开关活塞上设置有能够和所述定量腔连通的第二微通道。
  14. 根据权利要求13所述的环介导等温扩增芯片，其特征在于，所述反应区包括多个LAMP反应区和至少一个内标反应区，所述第一微通道的数量为多个，每个反应区分别对应一个所述第一微通道。
  15. 根据权利要求14所述的环介导等温扩增芯片，其特征在于，多个所述反应区和所述内标反应区沿x轴方向间隔排布，所述开关活塞能够沿x轴移动地插设于所述本体中。
  16. 根据权利要求14所述的环介导等温扩增芯片，其特征在于，所述开关活塞具有多个所述第一工作位置，当所述开关活塞在任一第一工作位置时，仅其中一个反应区通过相应的第一微通道和所述试剂区连通。
  17. 根据权利要求13所述的环介导等温扩增芯片，其特征在于，当所述开关活塞在第一工作位置时，所述定量腔通过第二微通道和所述试剂区相互连通。
  18. 根据权利要求13所述的环介导等温扩增芯片，其特征在于，所述开关活塞上设有对应所述试剂区的入口、对应所述反应区的出口及对应所述定量腔的第二入口。
  19. 根据权利要求18所述的环介导等温扩增芯片，其特征在于，所述第二入口为沿所述开关活塞的移动方向延伸的长孔。
  20. 根据权利要求11所述的环介导等温扩增芯片，其特征在于，所述本体上还设有与所述试剂区连通的通气口。
  21. 根据权利要求21所述的环介导等温扩增芯片，其特征在于，所述环介导等温扩增芯片还包括用于封闭所述通气口的密封膜。
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