WO2023193386A1 - 一种核酸检测微流控装置及其核酸检测方法 - Google Patents

一种核酸检测微流控装置及其核酸检测方法 Download PDF

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Publication number
WO2023193386A1
WO2023193386A1 PCT/CN2022/113382 CN2022113382W WO2023193386A1 WO 2023193386 A1 WO2023193386 A1 WO 2023193386A1 CN 2022113382 W CN2022113382 W CN 2022113382W WO 2023193386 A1 WO2023193386 A1 WO 2023193386A1
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WIPO (PCT)
Prior art keywords
cavity
reagent
housing
nucleic acid
test strip
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PCT/CN2022/113382
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English (en)
French (fr)
Inventor
张誉琳
胡雄浜
张国锋
黄建国
Original Assignee
厦门宝太生物科技股份有限公司
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Publication of WO2023193386A1 publication Critical patent/WO2023193386A1/zh

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    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L3/00—Containers or dishes for laboratory use, e.g. laboratory glassware; Droppers
    • B01L3/50—Containers for the purpose of retaining a material to be analysed, e.g. test tubes
    • B01L3/502—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures
    • B01L3/5027—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures by integrated microfluidic structures, i.e. dimensions of channels and chambers are such that surface tension forces are important, e.g. lab-on-a-chip
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
    • C12Q1/6806—Preparing nucleic acids for analysis, e.g. for polymerase chain reaction [PCR] assay
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
    • C12Q1/6844—Nucleic acid amplification reactions
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/70—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving virus or bacteriophage
    • C12Q1/701—Specific hybridization probes
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2200/00—Solutions for specific problems relating to chemical or physical laboratory apparatus
    • B01L2200/06—Fluid handling related problems
    • B01L2200/0689—Sealing
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2200/00—Solutions for specific problems relating to chemical or physical laboratory apparatus
    • B01L2200/10—Integrating sample preparation and analysis in single entity, e.g. lab-on-a-chip concept
    • 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
    • B01L2300/0867—Multiple inlets and one sample wells, e.g. mixing, dilution
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2400/00—Moving or stopping fluids
    • B01L2400/04—Moving fluids with specific forces or mechanical means
    • B01L2400/0475—Moving fluids with specific forces or mechanical means specific mechanical means and fluid pressure
    • B01L2400/0478—Moving fluids with specific forces or mechanical means specific mechanical means and fluid pressure pistons
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2400/00—Moving or stopping fluids
    • B01L2400/04—Moving fluids with specific forces or mechanical means
    • B01L2400/0475—Moving fluids with specific forces or mechanical means specific mechanical means and fluid pressure
    • B01L2400/0481—Moving fluids with specific forces or mechanical means specific mechanical means and fluid pressure squeezing of channels or chambers
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2400/00—Moving or stopping fluids
    • B01L2400/04—Moving fluids with specific forces or mechanical means
    • B01L2400/0475—Moving fluids with specific forces or mechanical means specific mechanical means and fluid pressure
    • B01L2400/0487—Moving fluids with specific forces or mechanical means specific mechanical means and fluid pressure fluid pressure, pneumatics
    • B01L2400/049—Moving fluids with specific forces or mechanical means specific mechanical means and fluid pressure fluid pressure, pneumatics vacuum
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
    • Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
    • Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
    • Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change

Definitions

  • the invention relates to the field of nucleic acid detection, and in particular to a nucleic acid detection microfluidic device and a nucleic acid detection method thereof.
  • corona virus disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) 2019 (COVID-19) is rapidly spreading and developing around the world. If a variety of infectious bacteria and viruses, including the new coronavirus, cannot be detected immediately, there will be risks of outbreaks and large-scale infections. With the recovery of the domestic and world economies, domestic and foreign personnel communications and business exchanges will become more frequent. The prevention and control of external imports poses greater challenges.
  • nucleic acid detection methods after collecting nasopharyngeal swab samples, the collected samples require specialized operators to perform sample inactivation, lysis, and magnetic beads in a biosafety level two or above laboratory. Enrichment extraction and final PCR amplification detection.
  • Traditional nucleic acid detection methods have high environmental requirements, complex operations, expensive equipment, and are time-consuming. They cannot perform rapid on-site testing, which greatly limits their application and promotion.
  • Reverse transcription-enhanced recombinase-dependent isothermal nucleic acid amplification technology combined with lateral flow chromatography test strips is a combination of reverse transcription-enhanced recombinase-dependent isothermal nucleic acid amplification technology (RT-eRDA technology) and lateral flow chromatography test strips
  • RT-eRDA technology reverse transcription-enhanced recombinase-dependent isothermal nucleic acid amplification technology
  • LFD lateral flow dipstick
  • RT-eRDA-LFD technology is based on RT-eRDA technology. It mainly uses a specific probe (nfo probe) with a 5' end labeled with a fluorescent group (usually FAM) and a 5' end labeled with biotin.
  • a double-labeled amplification product After incubating the primers at a constant temperature of 37°C to 42°C for 20 to 40 minutes, a double-labeled amplification product can be formed.
  • the amplification product to be detected flows to the binding area pre-coated with FAM antibody-gold nanoparticles using chromatography.
  • FAM antibody-gold nanoparticles combine with FAM in the double-labeled amplification product to form an immune complex.
  • the detection line (Test, T) is coated with a biotin antibody. When the immune complex diffuses to the detection line, the biotin in the double-labeled amplification product is captured by the biotin antibody and forms an immune complex visible to the naked eye.
  • the quality control line (Control, C) is coated with a secondary antibody, which can bind to the FAM antibody-gold nanoparticles and cause a color reaction and indicate the detection result.
  • this method Compared with traditional nucleic acid detection methods, this method has the characteristics of rapid response, intuitive judgment of detection results, and no need for special equipment. It is especially suitable for application scenarios such as low-resource areas, POCT, and on-site rapid detection. However, this method still has problems such as cumbersome operating steps, poor biological safety, low sensitivity, and low degree of integration.
  • Chinese patent CN 111733288 A discloses a nucleic acid detection method and its device and its application in COVID-19 detection.
  • RT-RAA-LFD technology is used to establish a microfluidic chip for rapid detection of the new coronavirus, and through specificity, sensitivity, practicality Sample analysis can be used for clinical on-site testing, providing a new sensitive and reliable method for instant detection of new coronavirus.
  • the internal structure of the device is fixed grooves and interconnected, which makes it impossible to pre-fill and isolate materials, such as buffers, freeze-dried reagents, etc., so all sample addition operations are dependent on the user, which is relatively cumbersome.
  • this invention is more inclined to be a conceptual device for experiment rather than industrialization.
  • Chinese patent CN 112760193 A discloses a nucleic acid extraction/detection device, which pre-embeds lysis solution, cleaning solution, eluent and nucleic acid amplification solution in different sealed and isolated areas of the device, and then simply rotates and squeezes the device The operation allows the relevant liquids to pass through the nucleic acid adsorption area in sequence, and the sample lysis, nucleic acid purification and nucleic acid amplification detection can be gradually realized, truly realizing the "sample in - result out" requirement in rapid nucleic acid on-site detection.
  • this device is a disposable sealed device, it can also effectively reduce and avoid the problem of cross-contamination of amplification products.
  • this invention mainly uses an ejection needle to pierce the sealing film to achieve the flow and mixing of various materials.
  • the sealing film piercing method generally has problems of random diameter and poor consistency after puncture, which can easily lead to The liquid mixing ratio cannot be reliably controlled, thus affecting the reaction effect and detection results.
  • the present invention provides a nucleic acid detection microfluidic device, which is portable and easy to operate and used for nucleic acid detection.
  • the detection time is fast, the detection results are highly accurate, and the biological safety is high.
  • the invention also provides a nucleic acid detection microfluidic device. A nucleic acid detection method using the nucleic acid detection microfluidic device.
  • the nucleic acid detection microfluidic device of the present invention includes a shell.
  • a lower piston cavity is provided inside the shell.
  • the piston in the lower piston cavity is connected to the lower piston rod.
  • the upper parts of the lower piston cavity are connected to each other.
  • the first reagent cavity and the second reagent cavity, the lower part of the lower piston cavity is connected to the reagent mixing part, one side of the reagent mixing part is connected to the test strip installation groove, and the test strip installation groove is inserted into Chromatography test paper strip, the chromatography test paper strip extends into the inside of the reagent mixing component, an opening is provided on one side of the housing, and a pressing device is provided in the opening, and the pressing device extends into the housing
  • the lower piston rod is connected internally and through gear transmission.
  • the top of the first reagent cavity is connected to the upper piston cavity
  • the piston in the upper piston cavity is connected to the upper piston rod
  • the upper piston cavity is connected to the first gas guide buffer cavity through the first air guide tube
  • the first gas guide buffer cavity is connected to the second gas guide buffer cavity
  • the second gas guide buffer cavity is connected to one side of the lower piston cavity
  • the top of the second reagent cavity passes through the second gas guide buffer cavity.
  • the air guide tube is connected to the first air guide buffer cavity
  • the pressing device is simultaneously connected to the upper piston rod through gear transmission.
  • two meshing gears are provided between the upper piston rod and the lower piston rod, which are a driving gear and a driven gear correspondingly installed in the gear installation groove.
  • the driving gear and the driven gear are
  • the driving gear and the second reagent chamber are respectively located on both sides of the first reagent chamber.
  • the driving gear and the driven gear are respectively connected to the inside of the housing through shaft rotation.
  • the upper piston rod The bottom of the lower piston rod meshes with the driving gear, the top of the lower piston rod meshes with the driven gear, and the driving gear meshes with the pressing device.
  • the pressing device includes a pressing device body, one side of the pressing device body extends downward to form a rack that meshes with the driving gear, a slide rail is provided inside the housing, and a sliding rail is provided on the pressing device body. There is a slider that can move up and down along the slide rail.
  • the pressing device body is provided with a first slot on the side away from the rack. There is a slot inside the housing that matches the first slot. protective lock, the upper edge of the pressing device body is arc-shaped and flush with the upper edge of the housing, the width of the upper edge of the pressing device body is greater than the width of the pressing device body, and is flush with the housing Consistent thickness.
  • the protection lock is located inside the housing, the protection lock includes a push rod, the upper edge of the push rod is flush with the upper edge of the housing, and a sliding block is provided at the lower end of the push rod, so One side of the sliding block is provided with a bayonet that matches the first slot, and the other side of the sliding block is provided with a snap-buckle mechanism.
  • the snap-buckle mechanism is arc-shaped, and the snap-buckle mechanism is located away from the One end of the sliding block is bent downward and extends in the direction of the bayonet.
  • the snap-lock mechanism is provided with latching teeth at one end away from the sliding block. There is a latching tooth inside the housing that is opposite to the latching tooth. Matching second and third card slots.
  • the lower piston rod includes a lower piston core, a first plug head is fastened around the lower piston core, a first plug body is provided on one side of the lower piston core, and a first plug body is provided on the top of the lower piston core.
  • the upper piston rod includes an upper piston core, a second plug head is fastened around the upper piston core, a second plug body is provided on one side of the upper piston core, and the bottom of the second plug body is provided with the
  • the drive gear is meshed with a rack structure, the second plug body is provided with a cavity structure, and the inside of the housing is provided with a limiting column that penetrates the cavity structure.
  • the reagent mixing component includes a mixing tank and a liquid storage tank, a flow limiting dam is provided between the lower part of the mixing tank and the lower part of the liquid storage tank, and the upper part of the mixing tank is connected with the upper part of the liquid storage tank.
  • the lower piston cavity is connected to the top of the mixing tank through a guide tube, and the top of the liquid storage tank is connected to the test strip installation groove;
  • the top of the test strip installation groove is connected to the top of the housing through a sealing plug.
  • the test strip installation groove is located close to the side of the housing.
  • the sealing plug is installed at the corner position of the top of the housing. .
  • the sealing plug includes a sealing plug body, the sealing plug body is ⁇ -shaped (door arch shape), a first buckle is provided at the lower part of one side of the sealing plug body, and the housing is provided with a first buckle inside.
  • the fourth snap slot is matched with the first buckle, and a sealing component is provided at the bottom of the other side of the sealing plug body.
  • the sealing component is inserted from the top of the test strip installation slot to form an interference fit.
  • the sealing The upper edge of the plug body is arc-shaped and flush with the upper edge of the housing.
  • the width of the upper edge of the sealing plug body is greater than the width of the sealing plug body and is consistent with the thickness of the housing.
  • the test strip installation groove is made of transparent material
  • the housing is provided with a transparent window corresponding to the detection result reading position of the chromatography test strip; the housing corresponds to the first reagent chamber.
  • a first sampling hole and a second sampling hole are respectively provided at the positions of the second reagent chamber and the first sampling hole and the second sampling hole are provided with matching first sampling plugs. and a second filling plug.
  • the invention also provides a method for nucleic acid detection using the above-mentioned nucleic acid detection microfluidic device, which includes the following steps:
  • the materials in the second reagent chamber and the first reagent chamber flow into the mixing tank through the lower piston cavity and the guide tube.
  • the first reagent and the second reagent are fully mixed in the mixing tank.
  • the liquid level in the mixing tank rises, and the material in the mixing tank overflows the current limiting dam and enters the liquid storage tank.
  • the chromatography test strip detects the material in the liquid storage tank. The color is developed through the CT line of the chromatography test strip and directly through the transparent window. Read the test results with the naked eye.
  • the present invention has the following effects:
  • the nucleic acid detection microfluidic device provided by the present invention has a highly integrated design that integrates various detection reaction materials and detection units into one body. It is low-priced, portable and easy to operate. It is used for nucleic acid detection, and the detection time is fast. The results are highly accurate and biosafety.
  • the inside of the kit (casing) of the present invention is sealed and isolated from the external space, and due to the formation of the vacuum cavity, a negative pressure environment is formed inside relative to the outside. Therefore, the biological components (such as nucleic acid amplification products) inside the kit are fully sealed in the internal cavity, eliminating the risk of overflow of aerosols and ensuring biological safety.
  • the present invention is suitable for nucleic acid detection based on isothermal nucleic acid amplification technology based on principles such as RT-eRDA-LFD. It is also suitable for detection that requires reliable control of multi-sample mixing, does not rely on special instruments to achieve detection, and ensures biological safety. system.
  • the second cavity of the present invention can be preloaded with biological components required for nucleic acid lysis, extraction, and amplification. After taking a nasopharyngeal swab, the user can directly add the sampling liquid into the second cavity to achieve accounting lysis. And expand directly.
  • the detection unit is a test strip. After the amplification is completed, the device realizes the mixing of various materials and the triggering and control of the detection process through a self-driven method, and all components are sealed inside the negative pressure device, fully ensuring biological safety.
  • the present invention adopts a self-driven design based on the principle of air pressure compensation balance, which can ensure that each material in the cavity is fully driven out, thereby ensuring the mixing ratio and sequence; the present invention integrates test strips as detection units, which can achieve no external The instrument relies on the result output; the invention does not have high requirements for operation, is simple and easy to operate, and can be used in industrial production.
  • Figure 1 is a schematic diagram of the internal structure of a nucleic acid detection microfluidic device in an embodiment of the present invention
  • Figure 2 is a schematic structural diagram of the front cover in the embodiment of the present invention.
  • Figure 3 is a schematic structural diagram of the back cover in the embodiment of the present invention.
  • Figure 4 is a schematic structural diagram of a pressing device in an embodiment of the present invention.
  • Figure 5 is a schematic structural diagram of the protection lock in the embodiment of the present invention.
  • Figure 6 is an exploded structural view of the lower piston rod in the embodiment of the present invention.
  • Figure 7 is an exploded view of the structure of the upper piston rod in the embodiment of the present invention.
  • Figure 8 is a schematic structural diagram of the sealing plug in the embodiment of the present invention.
  • Figure 9 is a schematic diagram of the process of nucleic acid detection using a nucleic acid detection microfluidic device in an embodiment of the present invention.
  • a nucleic acid detection microfluidic device includes a shell.
  • the shell can be made in one piece, or it can have a shell structure with two sides symmetrically matching each other.
  • the front cover 1 and the back cover 2 are symmetrically provided with several sets of assembly pins 23 structures.
  • the front cover 1 and the back cover 2 are assembled to form a shell structure, and the corresponding assembly pins 23 are assembled and tightened.
  • Existing processes (such as locking or welding, etc.) form the internal cavity structure of the shell, and the mutual joint surfaces are sealed and leak-free. As long as the internal structure of the shell is realized and the seal is leak-free.
  • FIG. 1 there is a lower piston cavity 9 inside the housing.
  • the piston in the lower piston cavity 9 is connected to the lower piston rod 10.
  • the upper part of the lower piston cavity 9 is connected to the first reagent chamber 16 and the second reagent chamber respectively.
  • Body 17 the lower part of the lower piston cavity 9 is connected to the reagent mixing part, one side of the reagent mixing part is connected to the test strip installation slot 22, the chromatography test strip is inserted into the test strip installation slot 22, and the chromatography test strip extends Inside the reagent mixing component, an opening is provided on one side of the housing, and a pressing device 4 is provided in the opening. The pressing device 4 extends into the interior of the housing and is connected to the lower piston rod 10 through gear transmission.
  • the top of the first reagent chamber 16 is connected to the upper piston chamber 7.
  • the piston in the upper piston chamber 7 is connected to the upper piston rod 6.
  • the upper piston chamber 7 is connected to the first air guide buffer chamber 20 through the first air guide tube 21.
  • a gas guide buffer cavity 20 is connected with the second gas guide buffer cavity 15
  • the second gas guide buffer cavity 15 is connected with one side of the lower piston cavity 9
  • the top of the second reagent cavity 17 is connected to the second gas guide buffer cavity 15 through the second gas guide tube 19 .
  • the first air guide buffer cavity 20 is connected, and the pressing device 4 is connected to the piston rod 6 through gear transmission.
  • the driving gear 8 and the driven gear 11 are respectively located on both sides of the first reagent chamber 16.
  • the driving gear 8 and the driven gear 11 are respectively connected inside the housing through shaft rotation.
  • the bottom of the upper piston rod 6 meshes with the driving gear 8, and the lower piston rod 6 meshes with the driving gear 8.
  • the top of 10 is meshed with the driven gear 11, and the driving gear 8 is meshed with the pressing device 4.
  • the pressing device 4 includes a pressing device body 4.1.
  • One side of the pressing device body 4.1 extends downward to form a rack 4.4 that meshes with the driving gear 8.
  • a slide rail 18 is provided inside the housing.
  • the pressing device body 4.1 is provided with a slide rail that can move up and down along the slider. As long as the up and down movement is achieved, the pressing device body 4.1
  • the side away from the rack 4.4 is provided with a first slot 4.2.
  • the inside of the casing is provided with a protective lock 5 that matches the first slot 4.2.
  • the upper edge of the pressing device body 4.1 is arc-shaped and is aligned with the upper edge of the casing. Flush, the width of the upper edge of the pressing device body 4.1 is greater than the width of the pressing device body 4.1, and is consistent with the thickness of the shell.
  • One side of the pressing device body 4.1 extends downward to form a rack 4.4 that meshes with the driving gear 8. It is used to cooperate with the driving gear 8 and the driven gear 11.
  • the driving gear 8 drives the driven gear 11 to run.
  • the pressing device 4 is used to accept the pressing action and drive the driving gear 8 to move.
  • the slide block 4.3 and the slide rail 18 play a limiting role, causing the pressing device 4 to move downward and drive the upper piston rod 6 to move (take Figure 1 as an example, (moves to the right), the driving gear 8 simultaneously drives the driven gear 11, and the driven gear 11 drives the lower piston rod 10 to move (taking Figure 1 as an example, moving to the right).
  • the protection lock 5 is located inside the shell. As shown in Figure 5, the protection lock 5 includes a push rod 5.3. The upper edge of the push rod 5.3 is flush with the upper edge of the shell. The lower end of the push rod 5.3 is provided with a sliding block 5.4.
  • the sliding block 5.4 is One side is provided with a bayonet 5.1 matching the first bayonet 4.2, and the other side of the sliding block 5.4 is provided with a snap-buckle mechanism 5.2.
  • the snap-buckle mechanism 5.2 is arc-shaped, and the end of the snap-buckle mechanism 5.2 away from the sliding block 5.4 faces toward It is bent downward and extends in the direction of the bayonet 5.1.
  • the end of the snap mechanism 5.2 away from the sliding block 5.4 is provided with bayonet teeth 5.5.
  • the interior of the housing is provided with a second bayonet 28 and a third bayonet that match the bayonet 5.5. 29.
  • the protection lock 5 is used to lock the pressing device 4 so that it cannot be pressed before the detection is started.
  • the snap mechanism 5.2 is arc-shaped. One end of the snap mechanism 5.2 is connected to the sliding block 5.4, and the other end is bent downward and extends in the direction of the bayonet 5.1. There is a second slot inside the housing that matches the bayonet 5.5. 28 and the third card slot 29. The second card slot 28 is located on the left side of the third card slot 29.
  • the snap mechanism 5.2 is used to limit the stroke range of the protection lock 5 to prevent malfunction.
  • the push rod 5.3 is the operating surface, and the user uses the protection lock 5 by pushing the push rod 5.3.
  • the push rod 5.3 When the push rod 5.3 is pushed (take Figure 1 as an example, push to the right), the push rod 5.3 drives the sliding block 5.4 to slide along the inside of the housing, and the bayonet 5.1 is pushed into the first slot 4.2 of the pressing device 4 to form a lock. At this time, the latching teeth 5.5 are engaged with the third latching slot 29 .
  • the push rod 5.3 When the push rod 5.3 is pushed (take Figure 1 as an example, push to the left), the push rod 5.3 drives the sliding block 5.4 to slide along the inside of the housing, and the bayonet 5.1 is pushed out of the first slot 4.2 of the pressing device 4, unlocking, and pushing When the latching teeth 5.5 engage with the second latching groove 28, the bayonet 5.1 and the first latching groove 4.2 of the pressing device 4 are unlocked.
  • the lower piston rod 10 includes a lower piston core 10.2.
  • a first plug head 10.3 is fastened around the lower piston core 10.2.
  • a first plug body 10.5 is provided on one side of the lower piston core 10.2.
  • the first plug body 10.5 is The top is provided with a first rack structure 10.1 that meshes with the driven gear 11, and an annular convex structure is provided on the outer periphery of the lower piston core 10.2.
  • the picture in the lower left corner of Figure 6 is a cross-sectional view of the first plug head, and the inside of the first plug head 10.3
  • There is a first annular groove structure 10.4 that matches the annular protruding structure.
  • the lower piston core 10.2 and the first plug head 10.3 form a tight fitting structure to prevent the first plug head 10.3 from falling off during operation.
  • the material of the first plug head 10.3 is rubber material.
  • a limiting device for the lower piston rod 10 can be provided on the housing.
  • a limiting support roller may be provided at the lower part of the lower piston rod 10, or a cavity structure may be provided on the lower piston rod, and a stopper may be provided inside the housing that penetrates the cavity structure.
  • the limiting column supports and guides the lower piston rod 10 during its operation, as long as the limiting and guiding functions of the lower piston rod 10 can be realized.
  • the lower piston rod 10 is used to be assembled in the lower piston cavity 9 to form a sealing effect on the first reagent cavity 16, the second reagent cavity 17, the second gas guide buffer cavity 15 and the flow guide tube 12, blocking and isolating The above-mentioned cavity and its contents.
  • the upper piston rod 6 includes an upper piston core 6.2.
  • a second plug head 6.3 is fastened around the upper piston core 6.2.
  • a second plug body 6.5 is provided on one side of the upper piston core 6.2.
  • the second plug body 6.5 is The bottom is provided with a second rack structure 6.1 that meshes with the driving gear 8.
  • the second plug body 6.5 is provided with a cavity structure 6.6.
  • the inside of the housing is provided with a limiting column 25 that penetrates the cavity structure 6.6.
  • the upper piston core 6.2 is provided with an annular protruding structure on its outer periphery.
  • the figure in the lower left corner of Figure 7 is a cross-sectional view of the second plug head 6.3.
  • the inner circumference of the second plug head 6.3 is provided with a second annular concave structure that matches the annular protruding structure.
  • the groove structure 6.4, the upper piston core 6.2 and the second plug head 6.3 form a tight fitting structure to prevent the second plug head 6.3 from falling off during operation.
  • the second plug head 6.3 is made of rubber material, and the second plug body 6.5 is There is a cavity structure 6.6, and the inside of the housing is provided with a limiting column 25 that penetrates the cavity structure 6.6.
  • the limiting column 25 supports and guides the upper piston rod 6 during its operation.
  • the upper piston rod 6 is used to be assembled in the upper piston cavity 7 to form a sealing effect on the first reagent cavity 16 and the first air conduit 21 to block and isolate the first reagent cavity 16 and the materials in it.
  • the reagent mixing component includes a mixing tank 24 and a liquid storage tank 14. There is a flow limiting dam 13 between the lower part of the mixing tank 24 and the lower part of the liquid storage tank 14. The upper part of the mixing tank 24 is connected with the upper part of the liquid storage tank 14. The lower piston cavity 9 The top of the mixing tank 24 is connected through the guide tube 12 , and the top of the liquid storage tank 14 is connected with the test strip installation slot 22 .
  • the top of the test strip installation groove 22 is connected to the top of the housing through the sealing plug 3.
  • the test strip installation groove 22 is located close to the side of the housing, and the sealing plug 3 is installed at the corner position of the top of the housing.
  • the sealing plug 3 includes a sealing plug body 3.3.
  • the sealing plug body 3.3 is in the shape of ⁇ .
  • the lower part of one side of the sealing plug body 3.3 is provided with a first buckle 3.2.
  • the inside of the shell can be provided with a first buckle 3.2.
  • the fourth matching slot is provided with a sealing component 3.1 at the bottom of the other side of the sealing plug body 3.3.
  • the sealing component 3.1 is inserted from the top of the test strip installation slot 22 to form an interference fit.
  • the upper edge of the sealing plug body 3.3 is arc-shaped. And it is flush with the upper edge of the casing.
  • the width of the upper edge of the sealing plug body 3.3 is greater than the width of the sealing plug body 3.3 and is consistent with the thickness of the casing.
  • the sealing plug 3 is installed from top to bottom at the corner position of the top of the housing.
  • the sealing component 3.1 is inserted from the top of the test strip installation slot 22 to form The interference fit forms a seal on the test strip installation slot 22.
  • a fourth slot that matches the first buckle 3.2 can be provided inside the housing.
  • the first buckle 3.2 is connected to the first buckle 3.2.
  • the four card slots are engaged and fixed to prevent the sealing plug 3 from falling off.
  • the test strip installation slot 22 and the shell are made of transparent material, or the shell is provided with a transparent window 30 at the position corresponding to the test result reading of the chromatography test strip.
  • This transparent window 30 is used to provide a complete and clear observation surface interface.
  • the transparent window 30 is designed with an outer edge for combining with the housing and achieving sealing (through welding or sealing rings, etc.).
  • the shell is provided with a first sampling hole 26 and a second sampling hole 27 respectively at positions corresponding to the first reagent chamber 16 and the second reagent chamber 17.
  • the first sampling hole 26 and the second sampling hole 27 are provided with Matching first and second sampling plugs.
  • the first sample filling plug and the second sample filling plug work together to achieve reliable sealing of the first reagent chamber 16 and the second reagent chamber 17 .
  • the first reagent chamber 16 can be pre-filled with required reagents during the manufacturing process, and can ensure the stability and preservation of the filled substances during transportation and turnover before subsequent delivery to the end user.
  • the method for nucleic acid detection using the above nucleic acid detection microfluidic device includes the following steps:
  • the chromatography test strip used in the present invention is a lateral flow dipstick (LFD); the amplification reagents and buffer reagents are conventional commercially available amplification reagents and buffer reagents in this field;
  • the incubation device can be an existing device capable of heating specific parts.
  • an incubation device with several holding mechanisms can be used to install several kit, and perform heating and incubation on the second chamber (the wireframe part in the lower left corner of the rightmost picture in the first row of Figure 9).
  • the heating and incubation conditions are: heating at a constant temperature of 37°C to 42°C for 20 ⁇ After the heating and incubation process is completed in 40 minutes, the incubation device preferably prompts the user through LED or buzzer.
  • the second reagent chamber and the second air guide buffer chamber are connected.
  • the material in the first reagent chamber flows into the mixing tank through the lower piston cavity and the guide tube.
  • the first reagent and the second reagent are fully mixed in the mixing tank.
  • the material in the mixing tank overflows the flow restriction.
  • the dam enters the liquid storage tank, and the chromatography test strip detects the materials in the liquid storage tank. The test result is read directly with the naked eye through the CT line color development of the chromatography test strip.
  • Figure 9 is a schematic diagram of the nucleic acid detection process of the nucleic acid detection microfluidic device of the present invention.
  • the upper left picture in the figure shows adding the sample
  • the upper middle picture shows the sealing plug
  • the upper right picture shows the second step inside the dotted box.
  • the reagent chamber is heated.
  • the right picture in the lower row shows the safety lock being opened.
  • the middle picture in the lower row shows the pressing device.
  • the left picture in the lower row shows the test results being read from one side (one side of the shell corresponds to the chromatography test strip test). There is a transparent observation window at the result reading position).
  • the application process of the present invention is as follows:
  • Kit (shell) assembly and preparation (1) Kit (shell) assembly and preparation:
  • the shell adopts a structure with two sides that cooperate with each other, namely the upper cover and the lower cover.
  • the upper cover and the lower cover are assembled opposite each other through existing processes (such as locking or welding, etc.) to form the main component of the reagent kit, forming a reagent.
  • the internal cavity structure of the box and the mutual joint surfaces are sealed without leakage.
  • the materials in the second reagent cavity and the first reagent cavity It will flow into the lower piston cavity one after another during the time when the pressure in the cavity is compensated until it reaches equilibrium, and then flows into the mixing tank through the guide tube.
  • the first reagent and the second reagent are fully mixed in the mixing tank;
  • the second reagent chamber is Designed to be opened first, the first reagent chamber is designed to be opened later, thereby ensuring that the material sample (such as buffer reagent) in the first reagent chamber and the material sample (such as nucleic acid amplification product) in the second reagent chamber are fully diluted and integration.
  • material sample such as buffer reagent
  • material sample such as nucleic acid amplification product
  • the inside of the kit is sealed and isolated from the external space, and due to the formation of a vacuum cavity, a negative pressure environment is formed inside relative to the outside. Therefore, the biological components (such as nucleic acid amplification products) inside the kit are fully sealed in the internal cavity, eliminating the risk of overflow of aerosols and ensuring biological safety.

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Abstract

本发明涉及核酸检测领域，尤其是涉及一种核酸检测微流控装置及其核酸检测方法。包括壳体，所述壳体内部设有下活塞腔体，所述下活塞腔体内活塞连接下活塞杆，所述下活塞腔体的上部分别连通第一试剂腔体和第二试剂腔体，所述下活塞腔体的下部连通试剂混合部件，所述试剂混合部件的一侧与试纸条安装槽连通，所述试纸条安装槽内插入层析试纸条，所述层析试纸条延伸入所述试剂混合部件内部，所述壳体一侧设有开口，所述开口设有按压装置，所述按压装置延伸入所述壳体内部并通过齿轮传动连接所述下活塞杆。本发明便携易操作，检测时间快，检测结果准确性高，生物安全性高。

Description

一种核酸检测微流控装置及其核酸检测方法 技术领域
本发明涉及一种核酸检测领域，尤其是涉及一种核酸检测微流控装置及其核酸检测方法。
 背景技术
现阶段，由严重急性呼吸综合征冠状病毒2 (severe acute respiratory syndrome coronavirus2,SARS-CoV-2)引起的新型冠状病毒肺炎(corona virus disease 2019 ,COVID‑19)在全球内快速蔓延及发展。如果不能对包括新冠病毒在内的多种传染性细菌、病毒进行即时检测，存在爆发和大规模传染风险，随着国内和世界经济的复苏，国内外的人员沟通、商务往来等会更加频繁，对外来输入的防控提出了更大的挑战。
然而，传统的核酸检测方法，在进行鼻咽拭子样本采集后，针对采集到的样本，需要在生物安全二级以上的实验室中，由专门的操作人员进行样本灭活，裂解，磁珠富集提取和最终的PCR扩增检测。传统核酸检测方法对环境要求高、操作复杂、设备昂贵且耗时长，不能进行现场快速检测，极大的限制了其应用及推广。
结合侧流层析试纸条的逆转录增强型重组酶依赖等温核酸扩增技术，是将逆转录增强型重组酶依赖等温核酸扩增技术(RT-eRDA技术)与侧流层析试纸条(Lateral flow dipstick，LFD)技术相结合，应用试纸条直接检测RT-eRDA扩增产物的一项技术。RT-eRDA-LFD技术以RT-eRDA技术为基础，主要利用一条5’末端标记荧光基团(通常为FAM)的特异性探针(nfo探针)和一条5’末端标记有生物素的反向引物，在恒温37℃～42℃下进行20～40分钟温育后即可形成具有双标记的扩增产物。当用试纸条进行检测时，待检扩增产物利用层析作用向预先包被有FAM抗体-纳米金颗粒的结合区流动。FAM抗体-纳米金颗粒与双标记的扩增产物中的FAM结合后形成免疫复合物。检测线(Test，T)处包被有生物素抗体，当免疫复合物扩散到检测线时，双标记的扩增产物中的生物素被生物素抗体捕获并形成肉眼可见的免疫复合物。质控线(Control，C)包被有二抗抗体，可与FAM抗体-纳米金颗粒结合并发生显色反应并指示检测结果。
相比于传统核酸检测方法，此方法以其反应快速、检测结果直观判别、无需特殊设备等特点，特别适用于低资源地区、POCT和现场快速检测等应用场景。然而这一方法仍然存在操作步骤繁琐、生物安全性差、灵敏度低、集成化程度低等问题。
中国专利CN 111733288 A公开了一种核酸检测的方法及其装置和在COVID-19检测中的应用，首次采用RT-RAA-LFD技术建立快速检测新冠病毒的微流控芯片，并通过特异性、灵敏度、实际样品分析，可用于临床现场检测，为新冠病毒的即时检测提供了一种灵敏、可靠的新方法。该装置的内部结构为固定沟槽，且相互联通，这导致其无法实现物料的预装填及隔离，如缓冲液、冻干试剂等，故所有加样操作均依赖于用户完成，较为繁琐。且由于其依赖于装置的放置方向实现检测流程，缺乏防呆防错机制，用户一旦意外误操作，如筹备或加样过程中翻倒等情况，将会导致检测失效，试剂盒报废。基于以上问题，该发明更偏向于实验用概念性装置，而非产业化定位。
中国专利CN 112760193 A公开了一种核酸提取/检测装置，通过预埋裂解液、清洗液、洗脱液和核酸扩增液在装置的不同密闭和隔离的区域，再通过对装置进行简单的旋转和挤压操作，使得相关的液体依次通过核酸吸附区域，就可以逐步实现样本裂解、核酸纯化和核酸扩增检测，真正实现了核酸现场快速检测中的“样本进-结果出”要求。同时由于本装置是一次性密闭装置，因此也可以有效的降低和避免扩增产物交叉污染的问题。但该发明主要采用顶针刺破封膜的方式，实现各物料的流动及混合，在实际产业化经验中，封膜刺破的方式普遍存在刺破后通路口径随机，一致性差的问题，易导致液体混合比例无法可靠控制，从而影响反应效果及检测结果。
因此，目前迫切需要开发出集成核酸提取、扩增、检测的集成化程度高、廉价、便携、操作便利、生物安全性高的一次性RT-eRDA-LFD核酸检测装置，能够通过简单的触发机制，即可实现在现场对核酸样本进行闭环的处理、检测及回收处置，从而摆脱对生物安全实验室的依赖，不管是在海关等防控外来输入压力较大的场景，还是在一些条件落后的偏远地区，都具有重要的意义。而目前在国内市场上能够实现上述效果的装置非常少，如何开发一种新的具有上述效果的核酸检测装置，对于下一阶段的疫情防控工作，将会带来巨大的帮助。
发明内容
针对现有技术存在的问题，本发明提供一种核酸检测微流控装置，便携易操作，用于核酸检测，检测时间快，检测结果准确性高，生物安全性高，本发明还提供一种采用该核酸检测微流控装置的核酸检测方法。
本发明所述的核酸检测微流控装置，包括壳体，所述壳体内部设有下活塞腔体，所述下活塞腔体内活塞连接下活塞杆，所述下活塞腔体的上部分别连通第一试剂腔体和第二试剂腔体，所述下活塞腔体的下部连通试剂混合部件，所述试剂混合部件的一侧与试纸条安装槽连通，所述试纸条安装槽内插入层析试纸条，所述层析试纸条延伸入所述试剂混合部件内部，所述壳体一侧设有开口，所述开口设有按压装置，所述按压装置延伸入所述壳体内部并通过齿轮传动连接所述下活塞杆。
优选地，所述第一试剂腔体顶部连通上活塞腔体，所述上活塞腔体内活塞连接上活塞杆，所述上活塞腔体通过第一导气管与第一导气缓冲腔体连通，所述第一导气缓冲腔体与第二导气缓冲腔体连通，所述第二导气缓冲腔体与所述下活塞腔体一侧连通，所述第二试剂腔体顶部通过第二导气管与所述第一导气缓冲腔体连通，所述按压装置同时通过齿轮传动连接所述上活塞杆。
优选地，所述上活塞杆与所述下活塞杆之间设有两个相啮合的齿轮，分别为对应安装在齿轮安装槽内的驱动齿轮和从动齿轮，所述驱动齿轮和所述从动齿轮、所述第二试剂腔体分别位于所述第一试剂腔体的两侧，所述驱动齿轮和所述从动齿轮分别通过轴转动连接在所述壳体内部，所述上活塞杆的底部与所述驱动齿轮相啮合，所述下活塞杆的顶部与所述从动齿轮相啮合，所述驱动齿轮与所述按压装置相啮合。
优选地，所述按压装置包括按压装置本体，所述按压装置本体一侧向下延伸形成与所述驱动齿轮相啮合的齿条，所述壳体内部设有滑轨，所述按压装置本体上设有可沿所述滑轨上下移动的滑块，所述按压装置本体远离所述齿条的一侧设有第一卡槽，所述壳体内部设有与所述第一卡槽相配合的保护锁，所述按压装置本体的上沿为弧形且与所述壳体上沿齐平，所述按压装置本体的上沿宽度大于所述按压装置本体的宽度，且与所述壳体厚度一致。
优选地，所述保护锁位于所述壳体内部，所述保护锁包括推动杆，所述推动杆的上沿与所述壳体上沿齐平，所述推动杆下端设有滑动块，所述滑动块的一侧设有与所述第一卡槽相配合的卡销，所述滑动块的另一侧设有弹扣机构，所述弹扣机构为弧形，所述弹扣机构远离所述滑动块的一端向下弯折且向所述卡销的方向延伸，所述弹扣机构远离所述滑动块的一端设有卡齿，所述壳体内部设有与所述卡齿相配合的第二卡槽和第三卡槽。
优选地，所述下活塞杆包括下活塞芯，所述下活塞芯外周紧固设置第一塞头，所述下活塞芯一侧设有第一塞体，所述第一塞体的顶部设有与所述从动齿轮相啮合的齿条结构；
所述上活塞杆包括上活塞芯，所述上活塞芯外周紧固设置第二塞头，所述上活塞芯一侧设有第二塞体，所述第二塞体的底部设有与所述驱动齿轮相啮合的齿条结构，所述第二塞体设有空腔结构，所述壳体内部设有贯穿所述空腔结构的限位柱。
优选地，所述试剂混合部件包括混合池和储液池，所述混合池下部和所述储液池下部之间设有限流坝，所述混合池的上部和所述储液池的上部连通，所述下活塞腔体通过导流管连接所述混合池顶部，所述储液池顶部与所述试纸条安装槽连通；
所述试纸条安装槽顶部通过密封塞与所述壳体顶部连接，所述试纸条安装槽位于靠近壳体的边侧设置，所述密封塞安装在所述壳体顶部的边角位置。
优选地，所述密封塞包括密封塞本体，所述密封塞本体为Π形状（门拱形状），所述密封塞本体一侧下部设有第一卡扣，所述壳体内部设有与所述第一卡扣相配合的第四卡槽，所述密封塞本体另一侧底部设有密封部件，所述密封部件从所述试纸条安装槽的顶部插入形成过盈配合，所述密封塞本体的上沿为弧形且与所述壳体上沿齐平，所述密封塞本体的上沿的宽度大于所述密封塞本体的宽度，且与所述壳体厚度一致。
优选地，所述试纸条安装槽采用透明材质，所述壳体对应所述层析试纸条的检测结果读取位置处设有透明视窗；所述壳体对应所述第一试剂腔体和所述第二试剂腔体的位置分别设有第一加样孔和第二加样孔，所述第一加样孔和所述第二加样孔设有相配合的第一加样塞和第二加样塞。
本发明还提供一种采用上述核酸检测微流控装置进行核酸检测的方法，包括以下步骤：
（1）将层析试纸条插入试纸条安装槽内，层析试纸条下端延伸入储液池内底部，安装密封塞使试纸条安装槽上端封口；
（2）将第一试剂由第一加样孔加入到第一试剂腔体内，将第二试剂由第二加样孔加入到第二试剂腔体内，分别采用第一加样塞和第二加样塞塞住第一加样孔和第二加样孔，此时壳体内部空间形成密封，与外部完全隔绝；第一试剂为缓冲试剂，第二试剂为病毒提取物样本和扩增试剂；
（3）对第二试剂腔体进行温育加热；温育结束后，推动推动杆，按压装置与保护锁解除锁定，按压按压装置使其下行，在按压装置下行的过程中，驱动齿轮和从动齿轮被带动运行，同步带动上活塞杆及下活塞杆朝壳体外部方向运动，导致上活塞腔体和下活塞腔体分别形成真空空腔，随着运动的发展，上活塞腔体的真空空腔和下活塞腔体的真空空腔同时与第一试剂腔体、第二试剂腔体、第一导气管、第二导气管、第一导气缓冲腔体及第二导气缓冲腔体形成联通，由于压强补偿特性，第二试剂腔体和第一试剂腔体的物料先后经下活塞腔体、导流管流入混合池内，第一试剂和第二试剂在混合池内充分混合，随着混合池内液位升高，混合池内物料漫过限流坝进入储液池内，层析试纸条对储液池内的物料进行检测，通过层析试纸条的CT线显色，通过透明视窗直接肉眼读取检测结果。
与现有技术相比，本发明具有以下效果：
（1）本发明提供的核酸检测微流控装置，高度集成化设计，将各类检测反应用途材料及检测单元集成于一体，价格低，便携易操作，用于核酸检测，检测时间快，检测结果准确性高，生物安全性高。
（2）本发明试剂盒（壳体）内部与外部空间密封隔绝，且由于真空空腔的形成，内部相对于外部形成负压环境。故试剂盒内部的生物成分(如核酸扩增产物)等，被充分密闭于内部腔体中，杜绝了气溶胶等的溢出风险，保证了生物安全性。
（3）本发明适用于基于RT-eRDA-LFD等原理的等温核酸扩增技术的核酸检测，也适用于需要可靠控制多样本混合，不依赖于专用仪器实现检测，并保证生物安全性的检测体系。
（4）本发明第二腔体可预装入核酸裂解、提取、扩增所需的生物成分，用户在鼻咽拭子取样后，可直接将取样液加入第二腔体中，实现核算裂解并直扩。检测单元为试纸条，在扩增完成后，装置通过自驱动方式实现各类物料的混合和检测流程触发及控制，且所有组分被密封于负压装置内部，充分保障了生物安全性。
（5）本发明采用基于气压补偿平衡原理的自驱动式设计，可确保腔体内的各物料被充分驱动流出，从而保证混合比例及顺序；本发明集成试纸条作为检测单元，可实现无外部仪器依赖的结果输出；本发明对操作的要求不高，简单易操作，能够产业化生产使用。
 附图说明
图1是本发明实施例中核酸检测微流控装置的内部结构示意图；
图2是本发明实施例中前盖的结构示意图；
图3是本发明实施例中后盖的结构示意图；
图4是本发明实施例中按压装置的结构示意图；
图5是本发明实施例中保护锁的结构示意图；
图6是本发明实施例中下活塞杆的结构分解图；
图7是本发明实施例中上活塞杆的结构分解图；
图8是本发明实施例中密封塞的结构示意图；
图9是本发明实施例中采用核酸检测微流控装置进行核酸检测的过程示意图；
图中：
1-前盖，2-后盖，3-密封塞，3.1-密封部件，3.2-第一卡扣，3.3-密封塞本体，4-按压装置，4.1-按压装置本体，4.2-第一卡槽，4.3-滑块，4.4-齿条，5-保护锁，5.1-卡销，5.2-弹扣机构，5.3-推动杆，5.4-滑动块，5.5-卡齿，6-上活塞杆，6.1-第二齿条结构，6.2-上活塞芯，6.3-第二塞头，6.4-第二环状凹槽结构，6.5-第二塞体，6.6-空腔结构，7-上活塞腔体，8-驱动齿轮，9-下活塞腔体，10-下活塞杆，10.1-第一齿条结构，10.2-下活塞芯，10.3-第一塞头，10.4-第一环状凹槽结构，10.5-第一塞体，11-从动齿轮，12-导流管，13-限流坝，14-储液池，15-第二导气缓冲腔体，16-第一试剂腔体，17-第二试剂腔体，18-滑轨，19-第二导气管，20-第一导气缓冲腔体，21-第一导气管，22-试纸条安装槽，23-组装销，24-混合池，25-限位柱，26-第一加样孔，27-第二加样孔，28-第二卡槽，29-第三卡槽，30-透明视窗。
具体实施方式
实施例
一种核酸检测微流控装置，包括壳体，壳体可以是一体制备而成，也可以是内部为两面相互对称配合的壳体结构，如图2和图3所示，分别为前盖1和后盖2，前盖1与后盖2的内部对称设有若干组组装销23结构，将前盖1与后盖2相向组装成型为壳体结构，将对应的组装销23组装拧紧，通过现有工艺(如锁扣或熔接等)，形成壳体内部腔体结构，且相互接合面密封无泄漏，只要实现壳体内部结构且密封无泄漏即可。
如图1所示，壳体内部设有下活塞腔体9，下活塞腔体9内活塞连接下活塞杆10，下活塞腔体9的上部分别连通第一试剂腔体16和第二试剂腔体17，下活塞腔体9的下部连通试剂混合部件，试剂混合部件的一侧与试纸条安装槽22连通，试纸条安装槽22内插入层析试纸条，层析试纸条延伸入试剂混合部件内部，壳体一侧设有开口，开口设有按压装置4，按压装置4延伸入壳体内部并通过齿轮传动连接下活塞杆10。
第一试剂腔体16顶部连通上活塞腔体7，上活塞腔体7内活塞连接上活塞杆6，上活塞腔体7通过第一导气管21与第一导气缓冲腔体20连通，第一导气缓冲腔体20与第二导气缓冲腔体15连通，第二导气缓冲腔体15与下活塞腔体9一侧连通，第二试剂腔体17顶部通过第二导气管19与第一导气缓冲腔体20连通，按压装置4同时通过齿轮传动连接上活塞杆6。
上活塞杆6与下活塞杆10之间设有两个相啮合的齿轮，分别为对应安装在齿轮安装槽内的驱动齿轮8和从动齿轮11，驱动齿轮8和从动齿轮11、第二试剂腔体17分别位于第一试剂腔体16的两侧，驱动齿轮8和从动齿轮11分别通过轴转动连接在壳体内部，上活塞杆6的底部与驱动齿轮8相啮合，下活塞杆10的顶部与从动齿轮11相啮合，驱动齿轮8与按压装置4相啮合。
如图4所示，按压装置4包括按压装置本体4.1，按压装置本体4.1一侧向下延伸形成与驱动齿轮8相啮合的齿条4.4，壳体内部设有滑轨18，按压装置本体4.1上设有可沿滑轨18上下移动的滑块4.3，或壳体内部设有滑块，按压装置本体4.1上设有可沿滑块上下移动的滑轨，只要实现上下移动即可，按压装置本体4.1远离齿条4.4的一侧设有第一卡槽4.2，壳体内部设有与第一卡槽4.2相配合的保护锁5，按压装置本体4.1的上沿为弧形且与壳体上沿齐平，按压装置本体4.1的上沿宽度大于按压装置本体4.1的宽度，且与壳体厚度一致。按压装置本体4.1一侧向下延伸形成与驱动齿轮8相啮合的齿条4.4，用来与驱动齿轮8和从动齿轮11配合，在按压装置4被按下时带动驱动齿轮8运行，驱动齿轮8带动从动齿轮11运行。按压装置4用于接受下压动作，带动驱动齿轮8动作，滑块4.3和滑轨18起到限位作用，使按压装置4向下运动，驱动上活塞杆6运动（以图1为例，向右运动），驱动齿轮同8时带动从动齿轮11，从动齿轮11带动下活塞杆10运动（以图1为例，向右运动）。
保护锁5位于壳体内部，如图5所示，保护锁5包括推动杆5.3，推动杆5.3的上沿与壳体上沿齐平，推动杆5.3下端设有滑动块5.4，滑动块5.4的一侧设有与第一卡槽4.2相配合的卡销5.1，滑动块5.4的另一侧设有弹扣机构5.2，弹扣机构5.2为弧形，弹扣机构5.2远离滑动块5.4的一端向下弯折且向卡销5.1的方向延伸，弹扣机构5.2远离滑动块5.4的一端设有卡齿5.5，壳体内部设有与卡齿5.5相配合的第二卡槽28和第三卡槽29。
保护锁5用于在检测启动前，锁定按压装置4使其无法被按下。弹扣机构5.2为弧形，弹扣机构5.2一端与滑动块5.4连接，另一端向下弯折且向卡销5.1的方向延伸，壳体内部设有与卡齿5.5相配合的第二卡槽28和第三卡槽29，第二卡槽28位于第三卡槽29的左侧，弹扣机构5.2用于限定保护锁5的行程范围，以防误动作。推动杆5.3为操作面，用户通过推动此推动杆5.3使用保护锁5。当推动推动杆5.3（以图1为例，向右推），推动杆5.3带动滑动块5.4沿壳体内部滑动，卡销5.1被推入按压装置4的第一卡槽4.2中，形成锁定，此时卡齿5.5与第三卡槽29卡合。当推动推动杆5.3（以图1为例，向左推），推动杆5.3带动滑动块5.4沿壳体内部滑动，卡销5.1被推出按压装置4的第一卡槽4.2中，解除锁定，推至卡齿5.5与第二卡槽28卡合，卡销5.1与按压装置4的第一卡槽4.2解除锁定。
如图6所示，下活塞杆10包括下活塞芯10.2，下活塞芯10.2外周紧固设置第一塞头10.3，下活塞芯10.2一侧设有第一塞体10.5，第一塞体10.5的顶部设有与从动齿轮11相啮合的第一齿条结构10.1，在下活塞芯10.2外周设置环状凸起结构，图6左下角的图为第一塞头的剖视图，第一塞头10.3内周设有与环状凸起结构相配合的第一环状凹槽结构10.4，下活塞芯10.2与第一塞头10.3形成紧固配合的结构，防止在运行过程中第一塞头10.3脱落，第一塞头10.3的材质为橡胶材质。可在壳体上设置下活塞杆10的限位装置，比如在下活塞杆10的下部设限位支撑辊，或在下活塞杆上设空腔结构，在壳体内部设有贯穿该空腔结构的限位柱，限位柱在下活塞杆10运行过程中对下活塞杆10进行支撑和导向，只要能够实现下活塞杆10的限位和导向功能即可。下活塞杆10用于装配于下活塞腔体9内，对于第一试剂腔体16、第二试剂腔体17、第二导气缓冲腔体15和导流管12形成密封作用，阻断隔离上述腔体及其内的物质。
如图7所示，上活塞杆6包括上活塞芯6.2，上活塞芯6.2外周紧固设置第二塞头6.3，上活塞芯6.2一侧设有第二塞体6.5，第二塞体6.5的底部设有与驱动齿轮8相啮合的第二齿条结构6.1，第二塞体6.5设有空腔结构6.6，壳体内部设有贯穿空腔结构6.6的限位柱25。上活塞芯6.2外周设置环状凸起结构，图7左下角的图为第二塞头6.3的剖视图，第二塞头6.3内周设有与环状凸起结构相配合的第二环状凹槽结构6.4，上活塞芯6.2与第二塞头6.3形成紧固配合的结构，防止在运行过程中第二塞头6.3脱落，第二塞头6.3的材质为橡胶材质，第二塞体6.5设有空腔结构6.6，壳体内部设有贯穿空腔结构6.6的限位柱25，限位柱25在上活塞杆6运行过程中对上活塞杆6进行支撑和导向。上活塞杆6用于装配于上活塞腔体7内，对于第一试剂腔体16和第一导气管21形成密封作用，阻断隔离第一试剂腔体16及其内的物质。
试剂混合部件包括混合池24和储液池14，混合池24下部和储液池14下部之间设有限流坝13，混合池24的上部和储液池14的上部连通，下活塞腔体9通过导流管12连接混合池24顶部，储液池14顶部与试纸条安装槽22连通。
试纸条安装槽22顶部通过密封塞3与壳体顶部连接，试纸条安装槽22位于靠近壳体的边侧设置，密封塞3安装在壳体顶部的边角位置。
如图8所示，密封塞3包括密封塞本体3.3，密封塞本体3.3为Π形状，密封塞本体3.3一侧下部设有第一卡扣3.2，壳体内部可设有与第一卡扣3.2相配合的第四卡槽，密封塞本体3.3另一侧底部设有密封部件3.1，密封部件3.1从试纸条安装槽22的顶部插入形成过盈配合，密封塞本体3.3的上沿为弧形且与壳体上沿齐平，密封塞本体3.3的上沿的宽度大于密封塞本体3.3的宽度，且与壳体厚度一致。在层析试纸条装入试纸条安装槽22后，将密封塞3由上向下安装在壳体顶部的边角位置，此时密封部件3.1从试纸条安装槽22的顶部插入形成过盈配合对试纸条安装槽22形成密封，为了达到更好的密封效果，可以在壳体内部可设有与第一卡扣3.2相配合的第四卡槽，第一卡扣3.2与第四卡槽卡合固定，防止密封塞3脱落。
试纸条安装槽22和壳体均采用透明材质，或者壳体对应层析试纸条的检测结果读取位置处设有透明视窗30，此透明视窗30用于提供完整清晰的观察面接口给用户辩读检测结果，此透明视窗30设计有一圈外沿，用于与壳体相互结合并实现密封(可通过熔接或密封圈等方式)。壳体对应第一试剂腔体16和第二试剂腔体17的位置分别设有第一加样孔26和第二加样孔27，第一加样孔26和第二加样孔27设有相配合的第一加样塞和第二加样塞。第一加样塞和第二加样塞共同作用以实现对第一试剂腔体16和第二试剂腔体17的可靠密封。第一试剂腔体16可在在制造过程中预先灌注所需试剂，并在后继交付至终端用户之前的运输周转过程中，保障所灌注物质的稳定及保存。
采用上述核酸检测微流控装置进行核酸检测的方法，包括以下步骤：
（1）将层析试纸条插入试纸条安装槽内，层析试纸条下端延伸入储液池内底部，安装密封塞使试纸条安装槽上端封口；
（2）将第一试剂由第一加样孔加入到第一试剂腔体内，将第二试剂由第二加样孔加入到第二试剂腔体内，分别采用第一加样塞和第二加样塞塞住第一加样孔和第二加样孔，此时壳体内部空间形成密封，与外部完全隔绝；第一试剂为缓冲试剂，第二试剂为病毒提取物样本和扩增试剂；本发明采用的层析试纸条是侧流层析试纸条(Lateral flow dipstick，LFD)；扩增试剂和缓冲试剂均采用本领域常规市售扩增试剂和缓冲试剂；
（3）对第二试剂腔体进行温育加热；温育装置采用现有的能够对特定部位进行加热的装置即可，比如可以采用带若干个托载机构的温育装置，用于安装若干个试剂盒，并对第二腔体（如图9第一排最右边的图中左下角的线框部分）进行加热温育，加热温育条件是：恒温37℃～42℃下加热20～40分钟，待加热温育过程完成后，温育装置优选通过LED或蜂鸣，对用户进行提示。
温育结束后，推动推动杆，按压装置与保护锁解除锁定，按压按压装置使其下行，在按压装置下行的过程中，驱动齿轮和从动齿轮被带动运行，同步带动上活塞杆及下活塞杆朝壳体外部方向运动，导致上活塞腔体和下活塞腔体分别形成真空空腔，随着运动的发展，上活塞腔体的真空空腔和下活塞腔体的真空空腔同时与第一试剂腔体、第二试剂腔体、第一导气管、第二导气管、第一导气缓冲腔体及第二导气缓冲腔体形成联通，由于压强补偿特性，第二试剂腔体和第一试剂腔体的物料先后经下活塞腔体、导流管流入混合池内，第一试剂和第二试剂在混合池内充分混合，随着混合池内液位升高，混合池内物料漫过限流坝进入储液池内，层析试纸条对储液池内的物料进行检测，通过层析试纸条的CT线显色，直接肉眼读取检测结果。
图9是本发明核酸检测微流控装置进行核酸检测的过程示意图，图中上排左图为加入样本，上排中间图为盖上密封塞，上排右图为对虚线框内部的第二试剂腔体进行加热，下排右图为拨开安全锁，下排中间图为按下按压装置，下排左图为从一侧读取检测结果（壳体一侧对应层析试纸条检测结果读取位置处设有透明观察窗）。
以基于RT-eRDA-LFD等温核酸扩增技术的核酸检测场景为例，本发明的应用过程如下：
（1）试剂盒（壳体）组装制备：
（a）壳体采用两面相互配合的结构，分别为上盖和下盖，将上盖和下盖通过现有工艺(如锁扣或熔接等)，相向组装成型为试剂盒主体部件，形成试剂盒内部腔体结构，且相互接合面密封无泄漏。
（b）将层析试纸条插入试纸条安装槽内，层析试纸条下端延伸入储液池内底部，安装密封塞使试纸条安装槽上端封口；
（c）将按压装置、驱动齿轮、从动齿轮、下活塞杆、下活塞腔体、上活塞杆和上活塞腔体，调整各部件齿位，使其相互咬合匹配，且按压装置的上沿与试剂盒上沿齐平，按压装置上的第一卡槽与保护锁上的卡销互锁，保护锁上的卡齿与壳体内部的第三卡槽卡合；
（2）测试过程：
（a）将第一试剂由第一加样孔加入到第一试剂腔体内，将第二试剂由第二加样孔加入到第二试剂腔体内，分别采用第一加样塞和第二加样塞塞住第一加样孔和第二加样孔，此时壳体内部空间形成密封，与外部完全隔绝；第一试剂为缓冲试剂，第二试剂为病毒提取物样本和扩增试剂；至此，试剂盒腔体内部空间形成密封，与外部完全隔绝。
（b）而后，将试剂盒放置于温育装置上，启动温育装置工作，对第二试剂腔体处实施温育。
（c）待温育结束后，推动推动杆（如图1所示向左推），推动杆带动滑动块沿壳体内部滑动，保护锁上的卡齿与第三卡槽不再卡合，推至保护锁上的卡齿与第二卡槽卡合，卡销被推出按压装置的第一卡槽中，解除锁定，向下按压按压装置。
（d）在按压装置下行的过程中，驱动齿轮和从动齿轮被带动运行，同步带动上活塞杆及下活塞杆朝壳体外部方向运动（如图1所示，向右运动），导致上活塞腔体和下活塞腔体分别形成真空空腔，随着运动的发展，上活塞腔体的真空空腔和下活塞腔体的真空空腔同时与第一试剂腔体、第二试剂腔体、第一导气管、第二导气管、第一导气缓冲腔体及第二导气缓冲腔体形成联通，此时由于压强补偿特性，第二试剂腔体和第一试剂腔体的物料，将在腔体内压力补偿直至平衡的时间内，先后流入下活塞腔体内，再通过导流管流入混合池内，第一试剂和第二试剂在混合池内充分混合；
（e）值得一提的是，本发明中，特别考虑了第二试剂腔体与第一试剂腔体在上活塞杆及下活塞杆的运动过程中的开启流动顺序，第二试剂腔体被设计为率先开启，第一试剂腔体被设计为后继开启，从而保证第一试剂腔体内的物质样本（如缓冲试剂）与第二试剂腔体内的物质样本(如核酸扩增产物)进行充分稀释及融合。类似的思路亦可被应用于其他使用场景中，通过调整上述相关结构的尺寸及排布位置予以达成，在此不再赘述。
（f）在上述混合物流入混合池内后，限流坝将阻挡混合液即时接触层析试纸条，而是随着混合池内的液位升高，漫过限流坝后，混合液进入储液池内方才与层析试纸条接触并触发检测。此设计的目的在于保证混合物流入混合池后，有充分的时间进一步均匀混合，然后进行检测，检测效果更准确。
（g）层析试纸条的CT线显色，用户直接肉眼读取检测结果。
（h）检测完成后，用户将试剂盒从温育装置上取下，装入试剂盒原包装封口袋内，进行回收处置。
由于试剂盒内部与外部空间密封隔绝，且由于真空空腔的形成，内部相对于外部形成负压环境。故试剂盒内部的生物成分(如核酸扩增产物)等，被充分密闭于内部腔体中，杜绝了气溶胶等的溢出风险，保证了生物安全性。
本发明所揭示的设计思路及各技术要点，并不局限于核酸检测。任何多样本混合检测的场景，均有机会从本发明所揭示的信息中获益。

Claims (10)

  1. 一种核酸检测微流控装置，其特征在于：包括壳体，所述壳体内部设有下活塞腔体，所述下活塞腔体内活塞连接下活塞杆，所述下活塞腔体的上部分别连通第一试剂腔体和第二试剂腔体，所述下活塞腔体的下部连通试剂混合部件，所述试剂混合部件的一侧与试纸条安装槽连通，所述试纸条安装槽内插入层析试纸条，所述层析试纸条延伸入所述试剂混合部件内部，所述壳体一侧设有开口，所述开口设有按压装置，所述按压装置延伸入所述壳体内部并通过齿轮传动连接所述下活塞杆。
  2. 根据权利要求1所述的核酸检测微流控装置，其特征在于：所述第一试剂腔体顶部连通上活塞腔体，所述上活塞腔体内活塞连接上活塞杆，所述上活塞腔体通过第一导气管与第一导气缓冲腔体连通，所述第一导气缓冲腔体与第二导气缓冲腔体连通，所述第二导气缓冲腔体与所述下活塞腔体一侧连通，所述第二试剂腔体顶部通过第二导气管与所述第一导气缓冲腔体连通，所述按压装置同时通过齿轮传动连接所述上活塞杆。
  3. 根据权利要求2所述的核酸检测微流控装置，其特征在于：所述上活塞杆与所述下活塞杆之间设有两个相啮合的齿轮，分别为对应安装在齿轮安装槽内的驱动齿轮和从动齿轮，所述驱动齿轮和所述从动齿轮、所述第二试剂腔体分别位于所述第一试剂腔体的两侧，所述驱动齿轮和所述从动齿轮分别通过轴转动连接在所述壳体内部，所述上活塞杆的底部与所述驱动齿轮相啮合，所述下活塞杆的顶部与所述从动齿轮相啮合，所述驱动齿轮与所述按压装置相啮合。
  4. 根据权利要求3所述的核酸检测微流控装置，其特征在于：所述按压装置包括按压装置本体，所述按压装置本体一侧向下延伸形成与所述驱动齿轮相啮合的齿条，所述壳体内部设有滑轨，所述按压装置本体上设有可沿所述滑轨上下移动的滑块，所述按压装置本体远离所述齿条的一侧设有第一卡槽，所述壳体内部设有与所述第一卡槽相配合的保护锁，所述按压装置本体的上沿为弧形且与所述壳体上沿齐平，所述按压装置本体的上沿宽度大于所述按压装置本体的宽度，且与所述壳体厚度一致。
  5. 根据权利要求4所述的核酸检测微流控装置，其特征在于：所述保护锁位于所述壳体内部，所述保护锁包括推动杆，所述推动杆的上沿与所述壳体上沿齐平，所述推动杆下端设有滑动块，所述滑动块的一侧设有与所述第一卡槽相配合的卡销，所述滑动块的另一侧设有弹扣机构，所述弹扣机构为弧形，所述弹扣机构远离所述滑动块的一端向下弯折且向所述卡销的方向延伸，所述弹扣机构远离所述滑动块的一端设有卡齿，所述壳体内部设有与所述卡齿相配合的第二卡槽和第三卡槽。
  6. 根据权利要求3所述的核酸检测微流控装置，其特征在于：所述下活塞杆包括下活塞芯，所述下活塞芯外周紧固设置第一塞头，所述下活塞芯一侧设有第一塞体，所述第一塞体的顶部设有与所述从动齿轮相啮合的齿条结构；
    所述上活塞杆包括上活塞芯，所述上活塞芯外周紧固设置第二塞头，所述上活塞芯一侧设有第二塞体，所述第二塞体的底部设有与所述驱动齿轮相啮合的齿条结构，所述第二塞体设有空腔结构，所述壳体内部设有贯穿所述空腔结构的限位柱。
  7. 根据权利要求1所述的核酸检测微流控装置，其特征在于：所述试剂混合部件包括混合池和储液池，所述混合池下部和所述储液池下部之间设有限流坝，所述混合池的上部和所述储液池的上部连通，所述下活塞腔体通过导流管连接所述混合池顶部，所述储液池顶部与所述试纸条安装槽连通；
    所述试纸条安装槽顶部通过密封塞与所述壳体顶部连接，所述试纸条安装槽位于靠近壳体的边侧设置，所述密封塞安装在所述壳体顶部的边角位置。
  8. 根据权利要求7所述的核酸检测微流控装置，其特征在于：所述密封塞包括密封塞本体，所述密封塞本体为Π形状，所述密封塞本体一侧下部设有第一卡扣，所述壳体内部设有与所述第一卡扣相配合的第四卡槽，所述密封塞本体另一侧底部设有密封部件，所述密封部件从所述试纸条安装槽的顶部插入形成过盈配合，所述密封塞本体的上沿为弧形且与所述壳体上沿齐平，所述密封塞本体的上沿的宽度大于所述密封塞本体的宽度，且与所述壳体厚度一致。
  9. 根据权利要求1所述的核酸检测微流控装置，其特征在于：所述试纸条安装槽采用透明材质，所述壳体对应所述层析试纸条的检测结果读取位置处设有透明视窗；所述壳体对应所述第一试剂腔体和所述第二试剂腔体的位置分别设有第一加样孔和第二加样孔，所述第一加样孔和所述第二加样孔设有相配合的第一加样塞和第二加样塞。
  10. 一种采用权利要求1-9任一所述的核酸检测微流控装置进行核酸检测的方法，其特征在于：包括以下步骤：
    （1）将层析试纸条插入试纸条安装槽内，层析试纸条下端延伸入储液池内底部，安装密封塞使试纸条安装槽上端封口；
    （2）将第一试剂由第一加样孔加入到第一试剂腔体内，将第二试剂由第二加样孔加入到第二试剂腔体内，分别采用第一加样塞和第二加样塞塞住第一加样孔和第二加样孔，此时壳体内部空间形成密封，与外部完全隔绝；第一试剂为缓冲试剂，第二试剂为病毒提取物样本和扩增试剂；
    （3）对第二试剂腔体进行温育加热；温育结束后，推动推动杆，按压装置与保护锁解除锁定，按压按压装置使其下行，在按压装置下行的过程中，驱动齿轮和从动齿轮被带动运行，同步带动上活塞杆及下活塞杆朝壳体外部方向运动，导致上活塞腔体和下活塞腔体分别形成真空空腔，随着运动的发展，上活塞腔体的真空空腔和下活塞腔体的真空空腔同时与第一试剂腔体、第二试剂腔体、第一导气管、第二导气管、第一导气缓冲腔体及第二导气缓冲腔体形成联通，由于压强补偿特性，第二试剂腔体和第一试剂腔体的物料先后经下活塞腔体、导流管流入混合池内，第一试剂和第二试剂在混合池内充分混合，随着混合池内液位升高，混合池内物料漫过限流坝进入储液池内，层析试纸条对储液池内的物料进行检测，通过层析试纸条的CT线显色，直接肉眼读取检测结果。
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