WO2022222209A1 - 一种自驱动微流控检测装置及其用途 - Google Patents

一种自驱动微流控检测装置及其用途 Download PDF

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WO2022222209A1
WO2022222209A1 PCT/CN2021/093862 CN2021093862W WO2022222209A1 WO 2022222209 A1 WO2022222209 A1 WO 2022222209A1 CN 2021093862 W CN2021093862 W CN 2021093862W WO 2022222209 A1 WO2022222209 A1 WO 2022222209A1
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primer
reverse
capillary
sample
self
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PCT/CN2021/093862
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English (en)
French (fr)
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尤其敏
周艳琼
帅金晓
林艺志
贾晓娟
林元奎
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杭州优思达生物技术有限公司
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Publication of WO2022222209A1 publication Critical patent/WO2022222209A1/zh

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    • 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
    • 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
    • C12Q1/6804—Nucleic acid analysis using immunogens
    • 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
    • C12Q1/6876—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
    • C12Q1/6888—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for detection or identification of organisms
    • C12Q1/689—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for detection or identification of organisms for bacteria
    • 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
    • C12Q1/6876—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
    • C12Q1/6888—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for detection or identification of organisms
    • C12Q1/6895—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for detection or identification of organisms for plants, fungi or algae
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/558—Immunoassay; Biospecific binding assay; Materials therefor using diffusion or migration of antigen or antibody
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/577—Immunoassay; Biospecific binding assay; Materials therefor involving monoclonal antibodies binding reaction mechanisms characterised by the use of monoclonal antibodies; monoclonal antibodies per se are classified with their corresponding antigens
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/58—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving labelled substances
    • G01N33/585—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving labelled substances with a particulate label, e.g. coloured latex
    • G01N33/587—Nanoparticles
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
    • G01N33/6893—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids related to diseases not provided for elsewhere
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N2333/00—Assays involving biological materials from specific organisms or of a specific nature
    • G01N2333/435—Assays involving biological materials from specific organisms or of a specific nature from animals; from humans
    • G01N2333/46—Assays involving biological materials from specific organisms or of a specific nature from animals; from humans from vertebrates
    • G01N2333/47—Assays involving proteins of known structure or function as defined in the subgroups
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N2800/00—Detection or diagnosis of diseases
    • G01N2800/34—Genitourinary disorders
    • G01N2800/347—Renal failures; Glomerular diseases; Tubulointerstitial diseases, e.g. nephritic syndrome, glomerulonephritis; Renovascular diseases, e.g. renal artery occlusion, nephropathy
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N2800/00—Detection or diagnosis of diseases
    • G01N2800/70—Mechanisms involved in disease identification
    • G01N2800/7095—Inflammation

Definitions

  • the invention relates to the field of biological detection, in particular to a self-driven microfluidic detection device and use thereof.
  • PCR instruments and constant temperature amplification instruments that apply the principle of nucleic acid amplification are gradually replacing traditional smear or microscopy methods in the field of pathogen and environmental microorganism detection; automated chemiluminescence platforms provide biochemical indicators, tumor markers and other detection items.
  • a sensitive, fast, high-throughput solution
  • POCT testing refers to a form of testing that analyzes immediately at the sampling site and obtains test results quickly.
  • the POCT nucleic acid detector integrates nucleic acid extraction, amplification, fluorescence detection and other functions into a miniaturized instrument, which can perform fast and accurate nucleic acid detection under non-laboratory conditions.
  • POCT testing equipment Compared with traditional large-scale equipment or standard laboratories, POCT testing equipment provides on-site testing methods for grassroots units, remote areas or other scenarios with insufficient medical conditions. At the same time, it also provides a convenient and fast solution for detection items that are more sensitive to detection time. Many POCT products such as small blood glucose meters, urine sugar test strip devices, etc. have begun to enter the home scene, and users complete the test in the form of self-test.
  • the market needs a low-cost, fast and easy-to-use detection device that can accurately control the sample addition and detection process, is safe and reliable, and can be used without professional training.
  • the present invention solves such a problem.
  • the purpose of the present invention is to provide a self-driven microfluidic detection device and its application.
  • the precise control of the sample adding and detection process is realized, the detection accuracy is improved, and the detection accuracy is improved.
  • Professional training can be quickly and easily operated, safe and reliable.
  • a self-driving microfluidic detection device comprising: an upper cover, a bottom plate fixed under the upper cover, a sandwich layer arranged between the upper cover and the bottom plate, a sample addition hole arranged on the upper cover, and a
  • the microfluidic diversion assembly is arranged on the interlayer and the position corresponds to the sample filling slot of the microfluidic diversion assembly, is arranged on the interlayer and is connected to the functional chamber of the microfluidic diversion assembly, and the result display of the detection result is displayed.
  • the microfluidic guide assembly includes: a capillary guide channel, an on-off connector that is connected between the capillary guide channels and controls the connection and disconnection of the capillary guide channels.
  • the capillary diversion channel is composed of a plurality of capillary diversion channel units; the capillary diversion channel unit includes: two parallel capillary tube diversion pieces formed between the capillary diversion pieces the diversion channel.
  • the distance between the two parallel capillary guide pieces is in the range of 0.01-2mm.
  • the attractive force between the liquid molecules in the guide channel is cohesive force
  • the attractive force between the liquid molecules in the guide channel and the capillary guide sheet is the adhesion force
  • Adhesion is greater than cohesion
  • the end portion of the capillary guide plate is provided with a bridge portion extending downward.
  • the capillary diversion channel is a tertiary capillary diversion channel
  • the functional chamber is a secondary functional chamber
  • the primary capillary diversion channel between the primary functional chambers is connected to the secondary capillary diversion channel between the on-off connector and the secondary functional chamber, and is connected to the secondary capillary diversion channel between the secondary functional chamber and the result display unit.
  • the primary function chamber is provided with a gap.
  • the on-off connector includes: a first buffer tank connected to the functional chamber, a second buffer tank connected to the first buffer tank and placed in the capillary diversion channel, and arranged in the first buffer tank.
  • the on-off connector includes: a connecting groove connected to the functional chamber, and a water-absorbing expansion member placed in the connecting groove and corresponding to the capillary diversion channel.
  • the capillary diversion channels are arranged to cross each other at a position below the sample addition hole, and the intersection is located below the center of the sample addition hole.
  • the aforementioned self-driven microfluidic detection device further comprises: a result reading window arranged on the upper cover and located above the result display part, a heating module attached to the bottom of the functional chamber and arranged in the bottom plate, connected to the heating module
  • the temperature control device of the module is connected to the power supply of the heating module and fixed in the bottom plate, connected to the power supply and fixed to the switch and indicator light in the bottom plate, set in the switch hole on the upper cover, and set in the indicator light window on the upper cover , the indicator light hole set on the mezzanine.
  • the aforementioned self-driven microfluidic detection device further comprises: buckles arranged on the upper cover and the bottom plate and used for assembly, and a result display member fixing component arranged on the upper cover and the interlayer, arranged on the upper cover and matched It is installed on the upper cover of the functional chamber of the functional chamber, and is fixed on the sample hole cover on the sample hole.
  • the device is pre-installed with nucleic acid detection reagents and used for biological nucleic acid detection.
  • a 4-channel self-driven microfluidic detection device is used.
  • the 4-channel self-driven microfluidic detection device is connected to the four functional chambers through the capillary diversion channel through the sample injection hole.
  • Each functional chamber corresponds to a chromatography test paper and a result. reading window;
  • the material of the capillary guide piece of the capillary guide channel is PE material, and the spacing of the capillary guide piece is 0.2mm;
  • the chromatography test paper used is nucleic acid immunochromatographic test paper
  • Step 1 Pre-install the new crown detection reagent system, the new crown B.1.1.7 detection reagent system, the A-fluid detection reagent system, and the B-fluid detection reagent system in 4 functional chambers;
  • the primer probes of the new crown detection reagent system include:
  • Step 2 collect the tester's throat swab sample; Step 3, immerse the swab head in the sampling solution and mix well; Step 4, place the device on the surface of the water platform, add all the sampling solution to the sample addition hole, and cover it for sample addition. hole cover; step 5, the reaction temperature is 50-65 °C, let stand for 10-30 minutes; step 6, immediately add the cleaning solution to the sample hole, cover the sample hole cover, and let stand for 5-15 minutes; step 7, Read the result.
  • a 2-channel self-driven microfluidic detection device is used.
  • the 2-channel self-driven microfluidic detection device is connected to the two functional chambers through the capillary diversion channel.
  • Each functional chamber corresponds to a chromatography test paper and a result. reading window;
  • the material of the capillary guide pieces of the capillary guide channel is glass, and the distance between the capillary guide pieces is 1mm;
  • the chromatography test paper used is nucleic acid immunochromatographic test paper
  • Step 1 pre-install the Escherichia coli detection reagent system and the Salmonella Enteritidis detection reagent system in two functional chambers;
  • the primer probes of the E. coli detection reagent system include:
  • the primer probes of Salmonella enterica detection reagent system include: forward peripheral primer: CGTGATGCTGAAAGTACCGA, reverse peripheral primer: GGCCGCCAAAACTTTCCTGA, forward cross primer: CCACCGCGTACGGACTTCACCGAAACACAAACGGGCAAG, reverse acceleration primer: AGATCTTTTAGCAATTGCTTCT, reverse probe 1: fluorescent marker -TGCCGCGCATACGGAACAG, reverse probe 2: fluorescent marker-CCACCGCGTACGGACTTCAC; step 2, use
  • a 2-channel self-driven microfluidic detection device is used.
  • the 2-channel self-driven microfluidic detection device is connected to the two functional chambers through the capillary diversion channel.
  • Each functional chamber corresponds to a chromatography test paper and a result. reading window;
  • the material of the capillary guide pieces of the capillary guide channel is glass, and the distance between the capillary guide pieces is 0.5mm;
  • the chromatography test paper used is nucleic acid immunochromatographic test paper
  • Step 1 pre-installing the amplification reagent system for specific detection of Salmonella Enteritidis and the quality control amplification reagent system with human GAPDH gene in two functional chambers;
  • the primer probes of the Salmonella Enteritidis amplification reagent system include: forward peripheral primer: CGTGATGCTGAAAGTACCGA, reverse peripheral primer: GGCCGCCAAAACTTTCCTGA, forward cross primer: CCACCGCGTACGGACTTCACCGAAACACAAACGGGCAAG, reverse acceleration primer: AGATCTTTTAGCAATTGCTTCT, reverse probe 1: fluorescent marker -TGCCGCGCATACGGAACAG, reverse probe 2: fluorescent marker-CCACCGCGTACGGACTTCAC;
  • the primer probes of the quality control amplification reagent system equipped with the human GAPDH gene include: forward peripheral primer: AGAACGGGAAGCTTGTCATC, reverse peripheral primer: CGAACATGGGGGCATCAG, forward cross Primer: CAGAGGGGGCAGAGATGAATCTTCCAGGAGCGAGATCC, reverse acceleration primer: ATCTTCCAGGAGCGAGATCCCAGAGGGGGCAGAGATGA, reverse probe 1: fluorescent marker-CAAAATCAAGTGGGGCGA, reverse probe 2:
  • the use of the aforementioned self-driven microfluidic detection device for the detection of microorganisms in urine samples includes the following contents:
  • a 4-channel self-driven microfluidic detection device is used.
  • the 4-channel self-driven microfluidic detection device is connected to the four functional chambers through the capillary diversion channel through the sample injection hole.
  • Each functional chamber corresponds to a chromatography test paper and a result. reading window;
  • the chromatography test paper used is nucleic acid immunochromatographic test paper
  • the material of the capillary guide pieces of the capillary guide channel is PE material, and the distance between the capillary guide pieces is 0.5mm;
  • Step 1 pre-install the Escherichia coli detection reagent system and the Salmonella Enteritidis detection reagent system in 4 functional chambers;
  • the primer probes of the Ureaplasma Detection Reagent System include:
  • the primer probes of the Staphylococcus aureus detection reagent system include: forward peripheral primer: CTGAATATGCAATGAAAGTAACTGA, reverse peripheral primer: TTTTTCTCTTTGCATATTATCGC, forward cross primer: GACAACGCTTCTTTATCATTTGTGACAAGAGCTAGAGTCGTTAGC, reverse acceleration primer: ATAATTTCTTCAAGTCGTGCCGC, reverse probe 1: fluorescence Marker-GTGATACCAGCATGAATCGGTTTA,
  • Reverse probe 1 fluorescent marker-GCCCAATCCCAAGCCGTCG
  • reverse probe 2 fluorescent marker-GAGGCCATTTACGCCCAATC
  • the primer probes of the Candida albicans detection reagent system include: forward peripheral primer: CGAGTTGCCCCAAGACATG, reverse peripheral primer: AATGACCGCTCTGAGTGATG, Forward cross primer: CAGGCCACAAACCCACCAAAGAGAATTGTCGAAAATCGCCCG, reverse acceleration primer: GTGCTCTAATGGGGCAATTTCCA, reverse probe 1: fluorescent marker-ATGCTGAGCCGGAGCCTTTA, reverse probe 2: fluorescent marker-CAGGCCACAAAACCCACCAAAGA;
  • Step 2 use a dry swab head Urine samples can be taken out when the top of the swab absorbs the sample; step 3, immerse the swab head in the sampling solution and mix well; step 4, place the device on the surface of the water platform, add all the sampling solution to the sample hole, Cover the sample addition hole; step 5, the reaction
  • the aforementioned self-driven microfluidic detection method for quantitatively detecting kidney markers includes the following contents:
  • a 4-channel self-driven microfluidic detection device is used.
  • the 4-channel self-driven microfluidic detection device is connected to the four functional chambers through the capillary diversion channel through the sample injection hole.
  • Each functional chamber corresponds to a chromatography test paper and a result. reading window;
  • the material of the capillary guide pieces of the capillary guide channel is PP material, and the distance between the capillary guide pieces is 0.4mm;
  • Step 1 prepare gold nanoparticles; Step 2, prepare gold-labeled RBP monoclonal antibody 1; Step 3, attach the NC membrane to the PVC bottom plate, and spray RBP monoclonal antibody 2 and goat antibody on each position of the NC membrane with a reciprocating film stripper
  • the mouse secondary antibody respectively the detection line (T line) and the quality control line (C line)
  • step 4 the cell fusion agent was added to the gold-labeled RBP monoclonal antibody solution; then sprayed on the binding pad of the glass fiber membrane , drying; step 5, assemble the sample pad of the glass cellulose film, the binding pad of the glass cellulose film, the NC film and the water-absorbing pad into a detection test strip, and adjust the sensitivity of the test strip;
  • step 6 assemble the detection test strip
  • the strip is assembled in the self-driven microfluidic detection device; in step seven, four gradients of normal saline are sequentially pre-installed in the four functional chambers; in step eight, the urine sample is added to the sample addition hole
  • the purposes of the aforementioned self-driven microfluidic detection device, applied to the detection of influenza A virus and influenza B virus, includes the following contents:
  • a 2-channel self-driven microfluidic detection device is used.
  • the 2-channel self-driven microfluidic detection device is connected to the two functional chambers through the capillary diversion channel.
  • Each functional chamber corresponds to a chromatography test paper and a result. Reading window; two T lines are set on the nucleic acid immunoassay test paper to display the detection results of influenza A and B respectively;
  • the chromatography test paper used is nucleic acid immunochromatographic test paper
  • the material of the capillary guide pieces of the capillary guide channel is glass, and the distance between the capillary guide pieces is 0.2mm;
  • Step 1 the influenza A virus detection reagent system and the influenza B virus detection reagent system are sequentially pre-installed in two functional chambers according to the components of the system;
  • the primer probes of the influenza A virus detection reagent system include: forward peripheral primer: CAGAGGGCAATGATGGATCA, reverse peripheral primer: ATCCCGACCAGTGAGTACC, forward cross primer: CCTCAGAATGAGTGCTGACCGTAAGTCGAAACCCAGGAAACG, reverse acceleration primer: CTTCCCTTTCAAAGTCATGCCCA, reverse probe 1: fluorescent marker- AGGAAAATGAGGTCTTCAATCTCAG, reverse probe 2: fluorescent marker-CCTCAGAATGAGTGCTGACCGT;
  • the primer probes of the B virus detection reagent system include: forward peripheral primer: ACTTACCAATGGGTGCTTAA, reverse peripheral primer: CGAAAAACAGAAAGGCAACAA, forward cross primer: GCATCCCATTGGAACATGTCTTCAAATTTAGTAACATTGAAGGCTCAG, reverse acceleration Primer: CCTCAGAAGATGGCTGGTCAGTTTTCATAACCTCTTGGTCTC, reverse probe 1: fluorescent marker-GTCTTCTTTTCCCAAAAGAAACTG, reverse probe
  • the device of the present invention adopts a microfluidic diversion design, the capillary diversion channels intersect with each other, and the intersecting structure can play the role of liquid storage and buffering, so the liquid balance in each flow direction can be realized; the structure of the capillary diversion sheet of the present invention
  • the cooperation with the material makes the adhesion greater than the cohesive force, and the liquid can infiltrate the contact surface and form a capillary action, thereby filling the space between the two capillary guide sheets and acting as a guide; thus realizing the precise control of liquid flow, While improving the accuracy, it avoids cross-contamination, and realizes the home self-inspection of nucleic acid detection;
  • the device of the invention does not need external power supply and water source; does not need professional laboratory environment; does not need professional training; in the case of improper operation, no harmful consequences such as electric shock and mechanical injury will occur, and the operation is simple and suitable for ordinary people to carry out home testing. As well as testing in some wild environments or under conditions of lack of professionals and equipment;
  • the device of the present invention can perform quantitative detection under the support of microfluidics and multiple detection;
  • the device of the present invention has high flexibility and can be applied to various fields such as infectious disease detection, pathogen typing detection, environmental microorganism detection, biochemical index detection, marker detection, drug detection and the like.
  • FIG. 1 is a top view of an embodiment of an upper cover of the present invention
  • Fig. 2 is a bottom view of an embodiment of an upper cover of the present invention
  • FIG. 3 is a side view of an embodiment of an upper cover of the present invention.
  • Figure 4 is a top view of an embodiment of the interlayer of the present invention.
  • Figure 5 is a cross-sectional view of an embodiment of the device of the present invention.
  • FIG. 6 is a schematic structural diagram of an embodiment of the on-off connector of the present invention.
  • FIG. 7 is a schematic structural diagram of the disconnected state of another embodiment of the on-off connector of the present invention.
  • FIG. 8 is a schematic structural diagram of the connection state of another embodiment of the on-off connector of the present invention.
  • FIG. 9 is a schematic structural diagram of an embodiment of a capillary guide sheet of the present invention.
  • FIG. 10 is a schematic structural diagram of another embodiment of the capillary guide sheet of the present invention.
  • a self-driven microfluidic detection device includes: an upper cover 1, a bottom plate 2 fixed under the upper cover 1, and disposed between the upper cover 1 and the bottom plate 2
  • the interlayer 3 is set on the sample injection hole 4 on the upper cover 1, and is connected to the microfluidic diversion assembly of the sample injection hole 4. It is set on the interlayer 3 and the position corresponds to the sample injection groove 7 of the microfluidic diversion assembly.
  • which is arranged on the interlayer 3 and connected to the functional chamber 8 of the microfluidic guide assembly, and displays the result display member of the detection result.
  • the result reading window 11 arranged on the upper cover 1 and located above the result display member is attached to the The heating module 10 at the bottom of the functional chamber 8 and arranged in the bottom plate 2, the temperature control device connected to the heating module 10, the power supply connected to the heating module 10 and fixed in the bottom plate 2, the power supply connected to the power supply and fixed in the bottom plate 2.
  • the switch 13, the indicator light are arranged on the switch hole 15 on the upper cover 1, the indicator light window 16 arranged on the upper cover 1, the indicator light hole 17 arranged on the interlayer 3, arranged on the upper cover 1 and the bottom plate 2 and used together
  • the result display member fixing components 20, 21 arranged on the upper cover 1 and the interlayer 3 are arranged on the upper cover 1 and matched with the functional chamber upper cover 14 of the functional chamber 8, and are fixed on the upper cover 1.
  • the result display part can be connected to different parts according to the detection type, and can be connected to the upper cover, mezzanine, bottom plate, etc. without limitation.
  • Reaction reagent It is the material used in the detection, which is pre-placed in the functional chamber. According to different detection items, the substances contained in the material and the pre-installation method can be combined in various ways.
  • the result display parts include: chromatography test paper 9, fluorescence detection instrument or test paper, visible dye detection test paper or device, electrical signal detector, etc., no matter the detection method or detection equipment is not limited, as long as it can connect the post-reaction reagents
  • the products showing the detection results are all suitable for the present invention; wherein the chromatography test paper 9 is used to present the detection results, the strip-shaped fiber chromatography material with the detection line and the quality control line fixed thereon is used as the stationary phase, the test liquid is the mobile phase, and the fluorescence is used as the mobile phase.
  • the labeled antibody or antigen is immobilized on the ligation pad, and the analyte is moved and captured on the chromatographic strip by capillary action for detection.
  • a functional chamber is a chamber that gives space for a physical or chemical reaction to occur; it is not limited by the type and object of detection.
  • the sample hole 4 and the sample hole cover 19 are located on the outer surface of one end of the top of the upper cover 1.
  • the inner diameter of the sample hole 4 matches the outer diameter of the cylindrical part below the sample hole cover 19.
  • the upper cover 14 of the functional chamber is located on the inner surface of the top of the upper cover 1 , and its shape and position match the functional chambers on the interlayer 3 .
  • the upper cover 14 of the functional chamber can cover the upper part of the functional chamber, so as to prevent the disorderly overflow of the liquid.
  • the result display member fixing components 20 and 21 are located on the inner surface of the top of the upper cover 1 and the upper surface of the interlayer 3, and are composed of result display member fixing protrusions. When the device is completely assembled, the result shows that the fixing protrusions of the piece play the role of fixing and holding up the immunological test strip.
  • the result reading window 11 is located on the top of the upper cover 1 and is made of transparent material. When the device is assembled, the result reading window 11 is located above the result display member. When using the device for testing, the test results can be read through the result reading window 11 .
  • the result reading window 11 can print pictures and texts to guide the result reading as needed.
  • Both the upper cover 1 and the bottom plate 2 are provided with buckles 18, the buckles 18 of the upper cover 1 are located on the inner surface of the side of the upper cover 1, and are used for assembling with the interlayer 3; the buckles 18 of the bottom plate 2 are used for and The upper cover 1 is fixedly assembled.
  • the sample adding groove 7 is located on the upper surface of the interlayer 3, and the sample adding groove 7 includes: the sample adding protrusions arranged at the edge position of the sample adding protrusions play the role of accommodating the liquid, and are formed between the sample adding protrusions and intersect with each other. like grooves.
  • the position of the sample addition groove 7 corresponds to the position of the sample addition hole 4
  • the sample addition groove of the sample addition groove 7 corresponds to the position of the microfluidic guide assembly of the upper cover 1 .
  • the functional chamber is where the reagents react or mix with the sample.
  • the microfluidic guide assembly is connected to the sample injection hole 4, the functional chamber, and the result display part.
  • the microfluidic diversion assembly includes: a capillary diversion channel 5 and an on-off connector 6 connected to the capillary diversion channel 5 and controlling the connection and disconnection of the capillary diversion channel 5 .
  • the capillary diversion channel 5 is composed of a plurality of capillary diversion channel units 51 ; the capillary diversion channel unit 51 includes: two parallel capillary diversion pieces 511 , and a diversion channel 512 formed between the capillary diversion pieces 511 .
  • the attractive force between the liquid molecules in the guide channel 512 is the cohesive force
  • the attractive force between the liquid molecules in the guide channel 512 and the capillary guide sheet 511 is the adhesion force; the adhesion force is greater than the cohesive force.
  • the material and surface texture of the capillary guide plate 511 are not limited, and can be adjusted according to the detected object; as an example, the material of the capillary guide plate 511 can be plastic, metal, glass, polymer, etc.;
  • the surface texture of the guide plate 511 can be a smooth surface, a frosted surface and a specific texture.
  • the distance between the two parallel capillary guide sheets 511 ranges from 0.01 to 2 mm. It should be noted that the examples here are not exhaustive, as long as the materials, textures, and spacings that can make the adhesion greater than the cohesive force fall within the protection scope of the present invention.
  • the end of the capillary guide piece 511 is provided with a bridge portion 5111 extending downward.
  • one end of the capillary guide piece 511 is provided with a bridge portion. 5111, the bridging part 5111 is bent downward, and the bridging part 5111 makes the end of the capillary guide plate 511 contact with the reaction material preloaded in the functional chamber.
  • two bridging portions 5111 are provided at both ends of the capillary guide sheet 511 , and the bridging portions 5111 extend downward, and have a U-shape as a whole.
  • the capillary diversion channels 5 are arranged to cross each other at a position below the sample addition hole 4 , and the intersection is located below the center of the sample addition hole 4 .
  • the device can also be composed of a multi-stage structure according to different detection requirements, the functional chamber and the capillary diversion channel: as shown in Figures 6 and 7, the specific structure is as follows.
  • the capillary diversion channel 5 is a tertiary capillary diversion channel 503
  • the functional chamber 8 is a secondary functional chamber 802
  • the tertiary capillary diversion channels 503 are respectively: connected to The primary capillary diversion channel 501 between the sample addition tank 7 and the primary functional chamber 801 is connected to the secondary capillary diversion channel 502 between the on-off connector 6 and the secondary functional chamber 802, and is connected to the second capillary diversion channel 502.
  • the tertiary capillary diversion channel 503 between the primary function chamber 802 and the chromatography test paper 9 .
  • the on-off connector 6 includes: a first buffer groove 601 connected to the primary functional chamber 801 , a second buffer groove 602 connected to the first buffer groove 601 and placed in the secondary capillary diversion channel 502 , a partition 603 disposed between the first buffer groove 601 and the second buffer groove 602 ; the height of the partition 603 is lower than the heights of the first buffer groove 601 and the second buffer groove 602 .
  • a gap 6011 is provided between the first buffer tank 601 and the primary function chamber 801 to guide the liquid to flow to the first buffer tank 601 .
  • the operation process is as follows: when the test sample is added from the sample injection hole 4, it is guided to the first-level functional chamber 801 through the first-level capillary diversion channel 501 for reaction; after the reaction, more liquid is added to make the liquid flow from the first-level The functional chamber 801 overflows into the first buffer tank 601.
  • the buffer groove 601 acts as a partition 603 between the first-level functional chamber 801 and the second buffer groove 602 , because the height of the partition 603 is lower than the height of the first buffer groove 601 and the second buffer groove 602 .
  • the liquid entering the first buffer tank 601 crosses the partition 603 and enters the second buffer tank 602, so as to realize the connection of the on-off connector 6, and enters the secondary functional chamber 802 through the secondary capillary channel 502 for reaction, and thereafter
  • the result is obtained by guiding the liquid to the chromatography strip 9 through the tertiary capillary channel 503 .
  • the on-off connector 6 includes: a connection groove 605 connected to the primary functional chamber 801 , and is placed in the connection groove 605 and corresponds to the water-absorbing expansion member under the secondary capillary diversion channel 502 .
  • the water-absorbing expansion member is a water-absorbing pad 604; it should be noted that the material and shape of the water-absorbing expansion member is not limited, as long as it can absorb water and expand to achieve on-off, it can be applied to the present invention.
  • the samples are evenly distributed to the primary functional chamber 801 for reaction after being added, and the liquid in the primary functional chamber 801 flows into the connection through the gap 6011 In the groove 605, the absorbent pad 604 absorbs water and expands. As shown in FIG. 8, it enters the connected state.
  • the absorbent pad 604 contacts the secondary capillary diversion channel 502, and the amplification products of the two reactions are diverted to the secondary capillary diversion channel 502 through the secondary capillary diversion channel 502.
  • the secondary functional chamber 802 is mixed in the secondary functional chamber 802, and then the secondary functional chamber 802 is connected to the nucleic acid immunochromatography test paper 9 through the tertiary capillary diversion channel 503.
  • the line captures the positive amplification products of the GAPDH system, and the T line captures the positive products of the Salmonella Enteritidis amplification system.
  • the first, second and third levels here are only for the purpose of clearly explaining the structure, and it is not necessary to have a second-level functional chamber and a third-level capillary diversion channel to use the on-off connector as described above.
  • the number of chambers and capillary channels is unlimited.
  • a connecting pipe can also be provided between the secondary capillary diversion channel 502 and the primary functional chamber 801, and the connecting pipe is connected to the primary functional chamber 801 obliquely or vertically.
  • the secondary capillary diversion channel 502 can be connected only after the amount of the two-stage capillary, so as to realize the on-off. It can also be switched on and off using hydraulic pressure.
  • the on-off can also be realized by mechanical devices, such as valves, etc.; the expansion member can also be expanded by the principle of thermal expansion, which will not be listed here.
  • the heating module 10 assembly includes: a heating module 10 attached to the bottom of the functional chamber and arranged in the bottom plate 2, a temperature control device connected to the heating module 10, a power supply connected to the heating module 10 and fixed in the bottom plate 2, connected to the heating module 10.
  • the power supply is fixed to the switch 13 and the indicator light in the bottom plate 2 , the switch hole 15 on the upper cover 1 , the indicator light window 16 on the upper cover 1 , and the indicator light hole 17 on the interlayer 3 .
  • Heating module 10 has the function of providing heating for the reaction, it is attached to the bottom of the functional chamber, and the temperature of the functional chamber is controlled by an electric heating and temperature control device.
  • Battery 12 The battery 12 is pre-installed in the space between the interlayer 3 and the bottom plate 2, and is connected to the heating module 10, the switch 13 and the indicator light by wires to provide power for the heating module 10.
  • Indicator light The indicator light plays the role of indicating whether the power is turned on. When the power is turned on, the indicator light is on.
  • the indicator window 16 is located at one end of the sample injection hole 4 of the upper cover 1 , and is made of light-transmitting material.
  • the portion of the indicator window 16 on the inner surface of the upper cover 1 has a raised structure for fixing the indicator light.
  • the switch hole 15 is located at one end of the sample injection hole 4 on the side of the upper cover 1 , and is used for installing the switch 13 .
  • the indicator light hole 17 is a circular hole for accommodating the indicator light and is located at one end of the sample injection hole 4 of the device; switch 13: is the power switch 13 of the device, and the current of the device can be turned on and off by opening and closing the switch 13, thereby controlling the detection reaction. conduct.
  • the device can adjust the number of channels according to different detection requirements.
  • the sample addition hole 4 is connected to two functional chambers through a capillary channel 5, and each functional chamber corresponds to a layer. Analysis test paper 9, a result reading window.
  • the sample addition hole 4 is connected to four functional chambers through the capillary diversion channel 5, and each functional chamber corresponds to a chromatography test paper 9 and a result reading window.
  • the sample addition hole 4 is connected to 6 functional chambers through the capillary diversion channel 5, and each functional chamber corresponds to a chromatography test paper 9 and a result reading window.
  • the device can choose to use each component according to the needs of the inspection items. For example, when the detection reaction does not require heating, the battery 12, the heating module 10, the switch 13, etc. may not be used to save manufacturing costs.
  • the operation steps of the device of the present invention include:
  • the detection device Place the detection device on the water platform, open the sample-adding hole cover 19, and add the sample into the sample-adding hole 4. Under the guidance of the microfluidic guide device, the liquid flows uniformly to each functional chamber.
  • the sample addition hole cover 19 is opened immediately, and buffer solution is added, so that the liquid inside the device flows from the functional chamber to the chromatography test paper 9 . Replace lid 1 and let stand for 5-15 minutes.
  • the detection accuracy of the device is verified through several detection applications of the device as follows.
  • Application 1 a method of applying a self-driving microfluidic nucleic acid detection device to nucleic acid detection of novel coronavirus/new coronavirus mutant strain/fluid A/B virus.
  • Test items new coronavirus, new coronavirus mutant strain B.1.1.7, influenza A virus, influenza B virus.
  • a 4-channel self-driven microfluidic detection device was used.
  • the sample addition hole 4 is connected to four functional chambers through the capillary diversion channel 5, and each functional chamber corresponds to a chromatography test paper 9 and a result reading window.
  • the material of the microfluidic guide device in this scheme is PE material, and the spacing of the guide plates is 0.2mm.
  • the adopted chromatography test paper 9 is nucleic acid immunochromatographic test paper 9 .
  • AMV reverse transcriptase AMV reverse transcriptase (NEB)
  • the pre-installed components in each functional chamber are: forward peripheral primer: 1-50 pmol, reverse peripheral primer: 1-50 pmol, forward cross primer: 10-500 pmol, reverse cross primer: 10-500 pmol, forward Probe: 1-100 pmol, reverse probe: 1-100 pmol, DNA polymerase: 5-50 U, reverse transcriptase: 0.5-8 U, dNTP: 10-300 nmol, BSA: 0.1-5 ⁇ g.
  • the materials of the four detection systems are pre-installed in the four functional chambers in sequence according to the components of the system.
  • Sampling solution composition 5-100mM Tris-HCl, 0.5-40 ⁇ M DTT, 0.1-1M Betaine, 0.1%-5% TritonX-100, 2-15mM MgSO 4 , pH 7-9@25°C.
  • Composition of cleaning solution 5-100mM Tris-HCl, 2-15mM NaCl.
  • step 4 After step 4 is completed, immediately add 0.2-1ml of the cleaning solution to the sample addition hole 4, cover the sample addition hole cover 19, and let it stand for 5-15 minutes;
  • the new coronavirus pseudovirus When the sample concentration was 100,000 copies/swab, the new coronavirus pseudovirus, the new coronavirus B.1.1.7 pseudovirus, the influenza A virus pseudovirus, and the influenza B virus pseudovirus were all detected, and the detection rate was 100%. .
  • the new coronavirus pseudovirus, new coronavirus B.1.1.7 pseudovirus, influenza A virus pseudovirus, and influenza B virus pseudovirus are all detected, and the detection rate is 100%. .
  • the new coronavirus pseudovirus, new coronavirus B.1.1.7 pseudovirus, influenza A virus pseudovirus, and influenza B virus pseudovirus are all detected, and the detection rate is 100%. .
  • the detection rate of new coronavirus pseudovirus is 80%; the detection rate of new coronavirus B.1.1.7 pseudovirus is 50%; the detection rate of influenza A virus pseudovirus is 30% %; the detection rate of influenza B virus pseudovirus was 40%;
  • the new coronavirus pseudovirus When the sample concentration was 10 copies/swab, the new coronavirus pseudovirus, new coronavirus B.1.1.7 pseudovirus, influenza A virus pseudovirus, and influenza B virus pseudovirus were not detected, and the detection rate was 0. %.
  • the new coronavirus pseudovirus, new coronavirus B.1.1.7 pseudovirus, influenza A virus pseudovirus, and influenza B virus pseudovirus were not detected, and the detection rate was 0. %.
  • the experimental results show that a self-driven microfluidic detection device can be used in the detection of four kinds of nucleic acids of new coronavirus, new coronavirus mutant B.1.1.7, influenza A virus, and influenza B virus, and the sensitivity is at or better than 1000 copies/ swab.
  • Application 2 a method for applying a self-driving microfluidic nucleic acid detection device to detecting food microorganisms.
  • Test items Escherichia coli, Salmonella Enteritidis.
  • a 2-channel self-driven microfluidic detection device is used, and the 2-channel self-driven microfluidic detection device is connected to the two functional chambers through the capillary diversion channel 4 through the sample injection hole 4.
  • Each functional chamber corresponds to a chromatography test strip 9 and a result reading window.
  • the microfluidic guide device of this scheme is made of glass, and the distance between guide plates is 1mm.
  • the adopted chromatography test paper 9 is nucleic acid immunochromatographic test paper 9 .
  • the ingredients pre-installed in each functional chamber are:
  • Forward peripheral primer 1-50pmol
  • Reverse peripheral primer 1-50pmol
  • Forward cross primer 10-500pmol
  • Reverse cross primer 10-500pmol
  • Forward probe 1-100pmol
  • Reverse probe 1-100 pmol
  • DNA polymerase 5-50 U
  • dNTP 10-300 nmol
  • BSA 0.1-5 ⁇ g.
  • the materials of the two detection systems are pre-installed in the two functional chambers in sequence according to the components of the systems.
  • Sampling solution composition 5-100mM Tris-HCl, 0.5-40 ⁇ M DTT, 0.1-50mg/ml BSA, 0.1-1M Betaine, 0.1%-5% TritonX-100, 2-15mM MgSO 4 , pH 7-9@25°C .
  • Composition of cleaning solution 5-100mM Tris-HCl, 2-15mM NaCl.
  • Step 1 Use a swab dipped in a small amount of sampling solution to wipe the surface of the food in a large area for 5-20 times;
  • Step 2 Immerse the swab head in 0.2-1ml of sampling solution, rotate or shake for 5-15 seconds;
  • Step 3 place the device on the water platform, add all the sampling liquid into the sample addition hole 4, and cover the sample addition hole cover 19;
  • Step 4 turn on the power switch 13, the reaction temperature is 50-75 ° C, and stand for 10-30 minutes;
  • Step 5 After Step 4 is completed, immediately add 0.2-1ml of the cleaning solution to the sample addition hole 4, cover the sample addition hole cover 19, and let it stand for 5-15 minutes;
  • Step 6 read the result.
  • the bacterial content of the swab samples collected from the three batches of food contaminated with Escherichia coli was: 13500 bacteria/swab, 6500 bacteria/swab, 16000 bacteria/swab; three The bacterial content of the swab samples collected from the food contaminated with Salmonella Enteritidis was: 3500 bacteria/swab, 1200 bacteria/swab, and 700 bacteria/swab.
  • test results show that the self-driven microfluidic detection device applied to the detection of food microorganisms can detect whether the food is contaminated by specific microorganisms within one hour, and the results are accurate, and the detection sensitivity is better than 1000 bacteria/swab.
  • Application 3 a method of applying a self-driven microfluidic detection device to the detection of microorganisms in urine samples
  • Test items Ureaplasma urealyticum, Staphylococcus aureus, Neisseria gonorrhoeae, Candida albicans
  • a 4-channel self-driven microfluidic nucleic acid detection device is adopted, and the 4-channel self-driven microfluidic detection device is connected to the four functional chambers through the capillary diversion channel 4 through the sample addition hole 4, and each functional chamber corresponds to a chromatography test paper 9. A result reading window.
  • the microfluidic guide device used in this scheme is made of PE material, and the distance between the guide plates is 0.5mm.
  • the adopted chromatography test paper 9 is nucleic acid immunochromatographic test paper 9 .
  • the primer probes are shown in Table 4 below:
  • the pre-installed components in each functional chamber are: forward peripheral primer: 1-50 pmol, reverse peripheral primer: 1-50 pmol, forward cross primer: 10-500 pmol, reverse cross primer: 10-500 pmol, forward Probe: 1-100 pmol, reverse probe: 1-100 pmol, DNA polymerase: 5-50 U, dNTP: 10-300 nmol, BSA: 0.1-5 ⁇ g.
  • the materials of the four detection systems are pre-installed in the four functional chambers in sequence according to the components of the system.
  • Sampling solution composition 5-100mM Tris-HCl, 0.5-40 ⁇ M DTT, 0.1 ⁇ 50mg/ml BSA, 0.1 ⁇ 1M Betaine, 0.1% ⁇ 5% TritonX-100, 2 ⁇ 15mM MgSO 4 , pH 7-10@25°C .
  • Composition of cleaning solution 5-100mM Tris-HCl, 2-15mM NaCl.
  • Step 1 use a dry swab head to take a urine sample, and when the top of the swab absorbs the sample, it can be taken out;
  • Step 2 Immerse the swab head in 0.2-1ml of sampling solution, turn to mix or shake for 5-15 seconds, and then let stand for 5-10 minutes;
  • Step 3 place the device on the water platform, add all the sampling liquid into the sample addition hole 4, and cover the sample addition hole cover 19;
  • Step 4 turn on the power switch 13 and let it stand for 30 minutes;
  • Step 5 After Step 4 is completed, immediately add 0.2-1ml of the cleaning solution to the sample addition hole 4, cover the sample addition hole cover 19, and let it stand for 5-15 minutes;
  • Step 6 read the result.
  • the urine containing Ureaplasma urealyticum, Staphylococcus aureus, Neisseria gonorrhoeae, Candida albicans was detected, and the pathogens in the urine were diluted and adjusted to 100000cfu/ml, 10000cfu/ml, 5000cfu/ml, 2000cfu/ml, 1000cfu/ml, 500cfu/ml, 200cfu/ml.
  • the detection rate of Ureaplasma urealyticum, Staphylococcus aureus, Neisseria gonorrhoeae was 100%, and the detection rate of Candida albicans was 60%.
  • the detection rate of Ureaplasma urealyticum was 100%
  • the detection rate of Staphylococcus aureus was 80%
  • the detection rate of Neisseria gonorrhoeae was 50%.
  • the detection rate of Candida albicans was 30%.
  • the detection rate of Ureaplasma urealyticum was 50%
  • the detection rate of Staphylococcus aureus was 40%
  • the detection rate of Neisseria gonorrhoeae was 20%
  • the detection rate of Candida albicans was not detected.
  • the detection rate of Ureaplasma was 10%, and Staphylococcus aureus, Neisseria gonorrhoeae and Candida albicans were not detected.
  • the experimental conclusion proves that the self-driven microfluidic detection device applied to the detection of microorganisms in urine samples can detect pathogens in urine.
  • the lower detection limits are: Ureaplasma 250cfu/ml, Staphylococcus aureus 500cfu/ml, Neisseria gonorrhoeae 500cfu/ml, Candida albicans 1000cfu/ml.
  • the fourth application is a method for quantitatively detecting kidney markers by applying a self-driven microfluidic detection device.
  • Test item retinol binding protein (RBP)
  • a 4-channel self-driven microfluidic detection device is adopted, and the 4-channel self-driven microfluidic detection device is connected to the four functional chambers through the capillary diversion channel 4 through the sample injection hole 4, and each functional chamber corresponds to a chromatography test paper 9 , a result reading window.
  • the guide vanes of the microfluidic guide device used in this scheme are made of PP material, and the spacing between the guide vanes is 0.4 mm.
  • Step 1 after heating the 0.01-0.1% gold chlorate solution to boiling, add 1% trisodium citrate under stirring, the volume ratio of the two is 10/1-100/1; when the solution completely turns into a transparent red Continue to boil for 5-30 minutes, and after stirring and cooling, the desired nano-gold is obtained.
  • Step 2 adjust the pH value of the nano-gold solution obtained in step 1 to 8-10 with potassium carbonate solution, add mouse anti-RBP monoclonal antibody 1, add the solution and mix evenly, the concentration is 20-50ng/ ⁇ l. Stir at room temperature for 5- After 40 minutes, add 10% calf serum solution in a volume ratio of 10/1-100/1 and stir for 20 minutes.
  • step 3 the solution prepared in step 2 is centrifuged at 10000-17000 rpm/min at low temperature for 10-30 minutes, and the supernatant is removed.
  • the pellet was resuspended to the original volume with borate buffer containing 5% BSA, centrifuged again in the same way to remove the supernatant, and then resuspended to 1/20 of the original volume by adding borate buffer containing 5% BSA.
  • Gold-labeled RBP monoclonal antibody 1 was obtained.
  • Step 4 Attach the NC film to the PVC bottom plate 2, and spray RBP monoclonal 2 and goat anti-mouse secondary antibody on different positions of the NC film with a reciprocating film marking machine, which are the detection line (T line) and the quality control line respectively. (Line C), dried at 37°C for 2 hours.
  • Step 5 add PVP, PEG, sucrose 0.02g/mL, 0.02g/mL, 0.5g/mL to the gold-labeled RBP monoclonal antibody solution obtained in step 3. Then spray it on the bonding pad of the glass fiber membrane, and dry it at 37°C for later use.
  • step 6 the sample pad, the binding pad, the NC membrane and the water-absorbing pad 604 of the glass cellulose membrane are assembled into a test strip.
  • the lower limit of sensitivity of the test strip was adjusted to 100 mg/L.
  • Step 7 Assemble the detection test strip in the self-driven microfluidic detection device.
  • Step 8 Prefill 0.2475mL, 0.225mL, 0.125mL, and 0mL of normal saline in sequence in the 4 functional chambers.
  • Step 9 add 0.6 mL of the urine sample into the sample-adding hole 4 of the detection device, and suck out the remaining sample in the hole after 1 minute. Under the control of a microfluidic flow-guiding device, access was made into 4 functional chambers, and the final volume of each chamber was 0.25 mL. Thus the samples were diluted 100-fold, 10-fold diluted, 2-fold diluted, and undiluted in the 4 functional chambers, respectively.
  • Step ten read the result.
  • Results 4 samples were tested, in which the RBP content was divided into healthy people, A sample concentration was 150mg/L, B sample 300mg/L, and people with renal inflammation, RBP concentration was C sample 1200mg/L and D sample 1600mg /L.
  • the self-driven microfluidic detection device of the present invention can be used to detect kidney markers and perform preliminary quantification, so as to provide preliminary judgment basis for kidney health.
  • Application five a method for applying a self-driving microfluidic nucleic acid detection device to the detection of A flow and B flow.
  • Test items influenza A virus, influenza B virus.
  • a 2-channel multi-stage self-driven microfluidic detection device is adopted.
  • the 2-channel self-driven microfluidic detection device is connected to two functional chambers through the capillary diversion channel 4 through the sample injection hole 4, and each functional chamber corresponds to a chromatography Test strip 9, a result reading window.
  • the sample After the sample enters from the sample addition hole 4, it enters the first-level functional chamber 801 through the first-level microfluidic diversion channel. , so that the volume of the added liquid exceeds the spiritual value of the on-off connector 6, so that the on-off connector 6 is activated so that the liquid enters the secondary functional chamber 802 through the secondary microfluidic diversion channel, and in the secondary functional chamber 802
  • the two reactants are thoroughly mixed, and then flow to the nucleic acid immunochromatography test paper 9 through the three-stage microfluidic diversion channel.
  • There are two T lines on the nucleic acid immunoassay test paper There are two T lines on the nucleic acid immunoassay test paper, which respectively display the detection results of influenza A and B.
  • the microfluidic guide device is made of glass, and the spacing between guide plates is 0.2mm.
  • the chromatographic test paper 9 used is a nucleic acid immunochromatographic test paper 9, and the detection results of A and B flow are presented on a test strip.
  • DNA polymerase Bst 3.0 DNA polymerase (NEB); Reverse transcriptase: AMV reverse transcriptase (NEB); the pre-installed components in each functional chamber are: Forward Peripheral Primer: 1-50pmol, Reverse Peripheral primer: 1-50pmol, forward cross primer: 10-500pmol, reverse cross primer: 10-500pmol, forward probe: 1-100pmol, reverse probe: 1-100pmol; DNA polymerase: 5-50U ; Reverse transcriptase: 0.5-8U; dNTP: 10-300 nmol; BSA: 0.1-5 ⁇ g.
  • the materials of the two detection systems are pre-installed in the two functional chambers in sequence according to the components of the systems.
  • Sampling solution composition 5-100mM Tris-HCl, 0.5-40 ⁇ M DTT, 0.1-1M Betaine, 0.1%-5% TritonX-100, 2-15mM MgSO 4 , pH 7-9@25°C.
  • Composition of cleaning solution 5-100mM Tris-HCl, 2-15mM NaCl.
  • step 4 After step 4 is completed, immediately add 0.2-1ml of the cleaning solution to the sample addition hole 4, cover the sample addition hole cover 19, and let it stand for 5-15 minutes;
  • the detection rate of influenza A virus pseudovirus was 85%, and the detection rate of influenza B virus pseudovirus was 90%.
  • the detection rate of influenza A virus pseudovirus was 30%; the detection rate of influenza B virus pseudovirus was 40%;
  • the detection rate was 0%.
  • the experimental results prove that a self-driven microfluidic detection device can be applied to the detection of four nucleic acids of influenza A virus and influenza B virus, and the results are finally presented on a test strip through the multi-stage microfluidic flow guiding device. Clear and clear, more intuitive.
  • Test item Salmonella Enteritidis
  • a 2-channel self-driven microfluidic detection device was used.
  • the microfluidic guide device of this scheme is made of glass, the distance between guide plates is 0.5mm, and the switch 13 device used is the water absorption pad 604 switch 13 device.
  • the water absorbing pad 604 in the switch 13 device cannot be connected because it absorbs less liquid and has a lower height than the secondary capillary channel 502.
  • the height of the absorbent pad 604 increases and is connected to the secondary capillary diversion channel 502, thereby connecting the secondary capillary diversion channel 502, and transporting the liquid to the secondary functional chamber 802, where The mixing takes place in the secondary functional chamber 802 .
  • the secondary functional chamber 802 is connected to the nucleic acid immunochromatographic test paper 9.
  • the C line of the chromatography test paper 9 captures the positive amplification product of the GAPDH system, and the T line captures the positive amplification product of the Salmonella Enteritidis amplification system. product.
  • the adopted chromatography test paper 9 is nucleic acid immunochromatographic test paper 9 .
  • the ingredients pre-installed in each functional chamber are:
  • Forward peripheral primer 1-50pmol; Reverse peripheral primer: 1-50pmol; Forward cross primer: 10-500pmol; Reverse cross primer: 10-500pmol; Forward probe: 1-100pmol; Reverse probe: 1-100 pmol; DNA polymerase: 5-50 U; dNTP: 10-300 nmol; BSA: 0.1-5 ⁇ g; the materials of the two detection systems are composed of the above components, and are sequentially pre-installed in two functional chambers.
  • Tris-HCl 0.5-40 ⁇ M DTT, 0.1-50mg/ml BSA, 0.1-1M Betaine, 0.1%-5% TritonX-100, 2-15mM MgSO 4 , pH 7-9@25°C; components of cleaning solution : 5-100 mM Tris-HCl, 2-15 mM NaCl.
  • Step 1 collect anal swabs
  • Step 2 Immerse the swab head in 0.2-1ml of sampling solution, rotate or shake to mix for 5-15 seconds, filter the sampling solution with a filter membrane, and filter out large-volume residues;
  • Step 3 place the device on the water platform, add all the sampling liquid into the sample addition hole 4, and cover the sample addition hole cover 19;
  • Step 4 turn on the power switch 13, the reaction temperature is 50-75 degrees, and stand for 10-30 minutes;
  • Step 5 After Step 4 is completed, immediately add 0.2-1ml of the cleaning solution to the sample addition hole 4, cover the sample addition hole cover 19, and let it stand for 5-15 minutes;
  • Step 6 read the result.
  • the above operation method collects samples and performs detection.
  • test results show that the self-driven microfluidic detection device is applied to detect Salmonella Enteritidis in anal swabs, and can stably detect samples with a sample concentration of 1000 bacteria/swab.
  • the device was demonstrated to be capable of detecting Salmonella Enteritidis in anal swabs.
  • the invention adopts the microfluidic diversion design, realizes the precise control of the liquid flow, the results are clearly presented, the reporting time is short, and accurate detection results can be obtained without professional training and expensive instruments. Improper operation of this product will not cause harmful consequences, and has extremely high safety. It is very suitable for ordinary people to conduct self-testing at home.
  • This product can detect samples such as urine, oropharyngeal swabs, anal swabs, and food scraping swabs, and has a very wide range of applications. Through home self-examination and choosing appropriate countermeasures, the anxiety of the masses can be alleviated, and unnecessary occupation of medical resources and crowd gathering can be avoided.

Abstract

一种自驱动微流控检测装置及其用途，装置包括：上盖(1)，固定于上盖(1)下的底板(2)，设置于上盖(1)和底板(2)之间的夹层(3)，设置于上盖(1)上的加样孔(4)，连接于加样孔(4)的微流控导流组件，设置于夹层(3)上且位置对应于微流控导流组件的加样槽(7)，设置于夹层(3)上并连接于微流控导流组件的功能腔室(8)，连接于微流控导流组件的层析试纸(9)；微流控导流组件包括：毛细管导流渠(5)，连接于毛细管导流渠(5)并控制毛细管导流渠(5)的连通和断开的通断连接器(6)；实现了加样和检测过程的精确控制，提高了检测准确性，无需专业培训即可快速简便的操作，安全可靠；可对多种样本进行检测，具有十分广泛的适用范围。

Description

一种自驱动微流控检测装置及其用途 技术领域
本发明涉及生物检测领域，特别是一种自驱动微流控检测装置及其用途。
背景技术
随着科学技术的发展，病原体、环境微生物、生化指标、肿瘤和器官标志物、毒品等领域的检测技术和设备得到了长足的发展。如应用核酸扩增原理的PCR仪、恒温扩增仪，在病原体、环境微生物检测领域正在逐渐取代传统的涂片或镜检的方法；自动化化学发光平台为生化指标、肿瘤标志物等检测项目提供了灵敏、快速、高通量的解决方案。
检测需求的扩大和检测场景丰富，使得POCT(point-of-care testing)式的检测成为近年来检测领域发展的主要方向之一。POCT检测是指在采样现场即刻进行分析，快速得到检验结果的一种检测形式。如POCT核酸检测仪，将核酸提取、扩增、荧光检测等功能全部整合在一台小型化的仪器上，可以在非实验室条件下进行快速、准确的核酸检测。
与传统的大型设备或标准实验室相比，POCT检测设备为广大基层单位，偏远地区或其他医疗条件不足的场景，提供了现场检测手段。同时，它也为对检测时间较为敏感的检测项目提供了便捷的快速方案。许多POCT产品如小型血糖仪，尿糖试纸装置等已经开始走进居家场景，由使用者以自检的形式完成检测。
许多情况下，人们希望能够通过自检的方式完成检测；而在一些野外环境或者缺乏专业人员、设备的情况下，人们更是迫切需要一种成本低廉，快速简单，结果准确，不需要专业培训即可使用的检测装置。该装置应具有广泛的场景适应性和使用安全性，能够在没有外部电源、水源的情况下进行检测。即使出现错误的操作，该装置亦不会对使用者产生危害性的后果。
目前的小型检测装置依然存在着诸多的问题：1，部分小型化检测仪器依然需要外部电源、水源，限制了应用的环境；2，目前小型化检测仪器对于老百姓来说，依然存在成本过高的问题；3，由于核酸检测往往需要进行核酸扩增或荧光信号的采集和分析，需要较为复杂的设备和专业技巧，目前国内居家自检的设备，仍不能实现核酸检测；4，目前的试纸类自检产品虽然满足了一定的要求，但是依然存在加样不精确，难以进行定量和多重检测困难等问题。5，目前的家庭检测装置缺乏灵活性，存在重复开发和资源浪费的问题。
市场需要一种成本低廉，快速简便，能够实现加样和检测过程的精确控制，安全可靠、无需经过专业培训即可使用的一次性检测装置，本发明解决这样的问题。
发明内容
为解决现有技术的不足，本发明的目的在于提供一种自驱动微流控检测装置及其用途，采用微流控技术，实现了加样和检测过程的精确控制，提高检测准确性，无需专业培训即可快速简便的操作，安全可靠。
为了实现上述目标，本发明采用如下的技术方案：
一种自驱动微流控检测装置，包括：上盖，固定于上盖下的底板，设置于上盖和底板之间的夹层，设置于上盖上的加样孔，连接于加样孔的微流控导流组件，设置于夹层上并位置对应于微流控导流组件的加样槽，设置于夹层上并连接于微流控导流组件的功能腔室，显示检测结果的结果显示件；微流控导流组件包括：毛细管导流渠，连接于毛细管导流渠之间并控制毛细管导流渠的联通和断开的通断连接器。
前述的一种自驱动微流控检测装置，毛细管导流渠由多个毛细管导流渠单元组成；毛细管导流渠单元包括：两片平行的毛细管导流片，形成于毛细管导流片之间的导流通道。
前述的一种自驱动微流控检测装置，两片平行的毛细管导流片之间的间距范围为：0.01-2mm。
前述的一种自驱动微流控检测装置，在导流通道内的液体分子之间的吸引力为内聚力，在导流通道内的液体分子与毛细管导流片之间的吸引力为附着力；附着力大于内聚力。
前述的一种自驱动微流控检测装置，毛细管导流片的端部设置有向下延伸的桥接部。
前述的一种自驱动微流控检测装置，毛细管导流渠为三级毛细管导流渠，功能腔室为二级功能腔室；三级毛细管导流渠分别是：连接于加样槽与一级功能腔室之间的一级毛细管导流渠，连接于通断连接器与二级功能腔室之间的二级毛细管导流渠，连接于二级功能腔室与结果显示件之间的三级毛细管导流渠。
前述的一种自驱动微流控检测装置，一级功能腔室设置有缺口。
前述的一种自驱动微流控检测装置，通断连接器包括：连接于功能腔室的第一缓冲槽，连接于第一缓冲槽并放置毛细管导流渠的第二缓冲槽，设置于第一缓冲槽与第二缓冲槽之间的隔断；隔断的高度低于第一缓冲槽、第二缓冲槽的高度。
前述的一种自驱动微流控检测装置，通断连接器包括：连接于功能腔室的连接槽，放置在连接槽内并对应于毛细管导流渠下的吸水膨胀件。
前述的一种自驱动微流控检测装置，毛细管导流渠在加样孔下方位置相互交叉设置，交叉点位于加样孔中心的下方。
前述的一种自驱动微流控检测装置，还包括：设置于上盖上并位于结果显示件上方的结果阅读窗口，贴合于功能腔室底部并设置于底板内的加热模块，连接于加热模块的温控装置，连接于加热模块并固定于底板内的电源，连接于电源并固定于底板内的开关、指示灯，设置于上盖上的开关孔，设置于上盖上的指示灯窗口，设置于夹层上的指示灯孔。
前述的一种自驱动微流控检测装置，还包括：设置于上盖、底板上并用于组装的卡扣，设置于上盖、夹层上的结果显示件固定组件，设置于上盖上并匹配于功能腔室的功能腔室上盖，固定于加样孔上的加样孔盖。
前述的一种自驱动微流控检测装置的用途，装置预装有核酸检测试剂并用于生物核酸检测。
前述的一种自驱动微流控检测装置的用途，用于核酸检测新冠/新冠突变株/甲流/乙流病毒核酸检测包括如下内容：
采用4通道自驱动微流控检测装置，4通道自驱动微流控检测装置为加样孔通过毛细管导流渠与四个功能腔室连接，每个功能腔室对应一条层析试纸，一个结果阅读窗；
毛细管导流渠的毛细管导流片的材质为PE材质，毛细管导流片的间距为0.2mm；
采用的层析试纸为核酸免疫层析试纸；
步骤一，将新冠检测试剂系统，新冠B.1.1.7检测试剂系统，甲流检测试剂系统，乙流检测试剂系统预装于4个功能腔室中；
新冠检测试剂系统的引物探针包括：
正向外围引物：GGCAGTCAAGCCTCTTCTC，反向外围引物：TCTGTCAAGCAGCAGCAAAG，正向交叉引物：TTCCCCTACTGCTGCCTGGAGTTCCTCATCACGTAGTCGC，反向加速引物：AAGAGCAGCATCACCG，反向探针1：荧光标记物-GAATTTCTTGAACTGTTGCG，反向探针2：荧光标记物-TTCCCCTACTGCTGCCTGGA；新冠B.1.1.7检测试剂系统的引物探针包括：正向外围引物：TTCTTTCACACGTGGTGT，反向外围引物：GACAGGGTTATCAAACCTCT，正向交叉引物：AGGTAAGAACAAGTCCTGAGTTGATTATTACCCTGACAAAGTTTTCAG，反向加速引物：GTACCATTGGTCCCAGA，反向探针1：荧光标记物-GTCCCAGAGATAGCATGG，反向探针2：荧光标记物-AGGTAAGAACAAGTCCTGAGTTGA；甲流检测试剂系统的引物探针包括：正向外围引物：CAGAGGGCAATGATGGATCA，反向外围引物：TCCCGACCAGTGAGTACC，正向交叉引物：CCTCAGAATGAGTGCTGACCGTAAGTCGAAACCCAGGAAACG，反向加速引物：CTTCCCTTTCAAAGTCATGCCCA，反向探针1：荧光标记物-AGGAAAATGAGGTCTTCAATCTCAG，反向探针2：荧光标记物-CCTCAGAATGAGTGCTGACCGT；乙流检测试剂系统的引物探针包括：正向外围引物： CTTACCAATGGGTGCTTAA，反向外围引物：CGAAAAACAGAAAGGCAACAA，正向交叉引物：ATCCCATTGGAACATGTCTTCAAATTTAGTAACATTGAAGGCTCAG，反向加速引物：CTCAGAAGATGGCTGGTCAGTTTTCATAACCTCTTGGTCTC，反向探针1：荧光标记物-GTCTTCTTTTCCCAAAAGAAACTG，反向探针2：荧光标记物-CAAGAGCAGTGCTCAAACAAATGA；
步骤二，采集测试者咽拭子样本；步骤三，将拭子头部浸入采样液中混匀；步骤四，将装置放置于水平台面，将采样液全部加入加样孔中，盖上加样孔盖；步骤五，反应温度50-65℃，静置10-30分钟；步骤六，随即将清洗液加入加样孔中，盖上加样孔盖，静置5-15分钟；步骤七，读取结果。
前述的一种自驱动微流控检测装置的用途，用于大肠杆菌、肠炎沙门氏菌检测包括如下内容：
采用2通道自驱动微流控检测装置，2通道自驱动微流控检测装置为加样孔通过毛细管导流渠与两个功能腔室连接，每个功能腔室对应一条层析试纸，一个结果阅读窗；
毛细管导流渠的毛细管导流片的材质为玻璃材质，毛细管导流片之间的间距为1mm；
采用的层析试纸为核酸免疫层析试纸；
步骤一，将大肠杆菌检测试剂系统，肠炎沙门氏菌检测试剂系统预装于2个功能腔室中；
大肠杆菌检测试剂系统的引物探针包括：
正向外围引物：ACCGTCAGGAAGCGGTAC，反向外围引物：TTTCACCCACTCTTCCTGGAT，正向交叉引物：AGACGGTTGGAGTTGGAGGAGTGCAGAACAGGCGGAAGTT，反向加速引物：GTCTTTCGCATCGTCAATCAAAA，反向探针1：荧光标记物-TTTTCGAACCGACCACCAACAC，反向探针2：荧光标记物-AGACGGTTGGAGTTGGAGGAGT；肠沙门氏菌检测试剂系统的引物探针包括：正向外围引物：CGTGATGCTGAAAGTACCGA，反向外围引物：GGCCGCCAAAACTTTCCTGA，正向交叉引物：CCACCGCGTACGGACTTCACCGAAACACAAACGGGCAAG，反向加速引物：AGATCTTTTAGCAATTGCTTCT，反向探针1：荧光标记物-TGCCGCGCATACGGAACAG，反向探针2：荧光标记物-CCACCGCGTACGGACTTCAC；步骤二，使用沾有少量采样液的拭子对待测物表面进行大范围擦拭5-20次；步骤三，将拭子头部浸入采样液中混匀；步骤四，将装置放置于水平台面，将采样液全部加入加样孔中，盖上加样孔盖；步骤五，反应温度50-75℃，静置10-30分钟；步骤六，随即将清洗液加入加样孔中，盖上加样孔盖，静置5-15分钟；步骤七，读取结果。
前述的一种自驱动微流控检测装置的用途，用于肛拭子中肠炎沙门氏菌检测包括如下内容：
采用2通道自驱动微流控检测装置，2通道自驱动微流控检测装置为加样孔通过毛细管导流渠与两个功能腔室连接，每个功能腔室对应一条层析试纸，一个结果阅读窗；
毛细管导流渠的毛细管导流片的材质为玻璃材质，毛细管导流片之间的间距为0.5mm；
采用的层析试纸为核酸免疫层析试纸；
步骤一，将特异性检测肠炎沙门氏菌的扩增试剂系统，装有人缘GAPDH基因的质控扩增试剂系统预装于2个功能腔室中；
肠炎沙门氏菌的扩增试剂系统的引物探针包括：正向外围引物：CGTGATGCTGAAAGTACCGA，反向外围引物：GGCCGCCAAAACTTTCCTGA，正向交叉引物：CCACCGCGTACGGACTTCACCGAAACACAAACGGGCAAG，反向加速引物：AGATCTTTTAGCAATTGCTTCT，反向探针1：荧光标记物-TGCCGCGCATACGGAACAG，反向探针2：荧光标记物-CCACCGCGTACGGACTTCAC；装有人缘GAPDH基因的质控扩增试剂系统的引物探针包括：正向外围引物：AGAACGGGAAGCTTGTCATC，反向外围引物：CGAACATGGGGGCATCAG，正向交叉引物：CAGAGGGGGCAGAGATGAATCTTCCAGGAGCGAGATCC，反向加速引物：ATCTTCCAGGAGCGAGATCCCAGAGGGGGCAGAGATGA，反向探针1：荧光标记物-CAAAATCAAGTGGGGCGA，反向探针2：荧光标记物-GGGAGCCAAAAGGGTC；步骤二，采集肛拭子；步骤三，将拭子头部浸入采样液中混匀，使用滤膜对采样液进行过滤，滤去残渣；步骤四，将装置放置于水平台面，将采样液全部加入加样孔中，盖上加样孔盖；步骤五，反应温度50-75℃，静置10-30分钟；步骤六，随即将清洗液加入加样孔中，盖上加样孔盖，静置5-15分钟；步骤七，读取结果。
前述的一种自驱动微流控检测装置的用途，用于尿液样本中微生物检测包括如下内容：
采用4通道自驱动微流控检测装置，4通道自驱动微流控检测装置为加样孔通过毛细管导流渠与四个功能腔室连接，每个功能腔室对应一条层析试纸，一个结果阅读窗；
采用的层析试纸为核酸免疫层析试纸；
毛细管导流渠的毛细管导流片的材质为PE材质，毛细管导流片之间的间距为0.5mm；
步骤一，将大肠杆菌检测试剂系统，肠炎沙门氏菌检测试剂系统预装于4个功能腔室中；
解脲支原体检测试剂系统的引物探针包括：
正向外围引物：GTGATTTAACTGTAGAACAAGAACA，反向外围引物：AGGACCACTATATTGTAGTAGTGC，正向交叉引物：GGCATGCGATATGAAACACCATAGATCTTTTTTGACCAGGATC，反向加速引物：ATTATGATTTTTAACTGGTTCTTC，反向探针1：荧光标记物-CACCATTTTTAATTACAGTAACT，反向探针2：荧光标记物-GGCATGCGATATGAAACACCA；金黄色葡萄球菌检测试剂系统的引物探针包括：正向外围引物：CTGAATATGCAATGAAAGTAACTGA，反向外围引物：TTTTTCTCTTTGCATATTATCGC，正向交叉引物：GACAACGCTTCTTTATCATTTGTGACAAGAGCTAGAGTCGTTAGC，反向加速引物：ATAATTTCTTCAAGTCGTGCCGC，反向探针1：荧光标记物-GTGATACCAGCATGAATCGGTTTA，反向探针2：荧光标记物-GACAACGCTTCTTTATCATTTGTGA；淋病奈瑟球菌检测试剂系统的引物探针包括：正向外围引物：GCTTTTAAATCCAATACCGTATT，反向外围引物：TTGAGTTCGATGGTGCTG，正向交叉引物：GAGGCCATTTACGCCCAATCAACAATAAAATATCCATCACCACTG，反向加速引物：GTGCCGTCAAGGGAAGGTTG，
反向探针1：荧光标记物-GCCCAATCCCAAGCCGTCG，反向探针2：荧光标记物-GAGGCCATTTACGCCCAATC；白色念球菌检测试剂系统的引物探针包括：正向外围引物：CGAGTTGCCCCAAGACATG，反向外围引物：AATGACCGCTCTGAGTGATG，正向交叉引物：CAGGCCACAAACCCACCAAAGAGAATTGTCGAAAATCGCCCG，反向加速引物：GTGCTCTAATGGGGCAATTTCCA，反向探针1：荧光标记物-ATGCTGAGCCGGAGCCTTTA，反向探针2：荧光标记物-CAGGCCACAAACCCACCAAAGA；步骤二，使用干燥的拭子头部沾取尿液样本，当拭子顶部吸收到样本即可取出；步骤三，将拭子头部浸入采样液中混匀；步骤四，将装置放置于水平台面，将采样液全部加入加样孔中，盖上加样孔盖；步骤五，反应温度50-75℃，静置10-30分钟；步骤六，随即将清洗液加入加样孔中，盖上加样孔盖，静置5-15分钟；步骤七，读取结果。
前述的一种自驱动微流控检测方法，用于定量检测肾脏标志物包括如下内容：
采用4通道自驱动微流控检测装置，4通道自驱动微流控检测装置为加样孔通过毛细管导流渠与四个功能腔室连接，每个功能腔室对应一条层析试纸，一个结果阅读窗；
毛细管导流渠的毛细管导流片的材质为PP材质，毛细管导流片之间的间距为0.4mm；
步骤一，制备纳米金；步骤二，制备金标RBP单克隆抗体1；步骤三，将NC膜附在PVC底板，用往复划膜仪在NC膜的各个位置喷RBP单克隆抗体2及羊抗鼠二抗，分别为检测线(T线)和质控线(C线)，烘干；步骤四，在金标RBP单克隆抗体溶液中加入细胞融合剂；然后喷于玻璃纤维膜的结合垫上，烘干；步骤五，将玻璃纤维素膜的样品垫、玻璃纤维素膜的结合垫、NC膜和吸水垫组装成为检测试纸条，调整试纸条的灵敏度；步骤六，将检测试纸条组装于自驱动微流控检测装置中；步骤七，在4个功能腔室中依次预装四个梯度的生理盐水；步骤八，将尿液样本加入检测装置的加样孔中，吸出多余的样本，在微流控导流装置的控制下，进入4个功能腔室中，样本在4个功能腔室中分别被稀释100倍，10倍稀释，2倍稀释，和不稀释。步骤九，读取结果。
前述的一种自驱动微流控检测装置的用途，应用于甲流病毒、乙流病毒检测包括如下内容：
采用2通道自驱动微流控检测装置，2通道自驱动微流控检测装置为加样孔通过毛细管导流渠与两个功能腔室连接，每个功能腔室对应一条层析试纸，一个结果阅读窗；在核酸免疫试纸上设置有两条T线，分别显示甲流和乙流的检测结果；
采用的层析试纸为核酸免疫层析试纸；
毛细管导流渠的毛细管导流片的材质为玻璃材质，毛细管导流片之间的间距为0.2mm；
步骤一，将甲流病毒检测试剂系统、将乙流病毒检测试剂系统，按照系统的成份组成，依次预装于两个功能腔室中；
甲流病毒检测试剂系统的引物探针包括：正向外围引物：CAGAGGGCAATGATGGATCA，反向外围引物：ATCCCGACCAGTGAGTACC，正向交叉引物：CCTCAGAATGAGTGCTGACCGTAAGTCGAAACCCAGGAAACG，反向加速引物：CTTCCCTTTCAAAGTCATGCCCA，反向探针1：荧光标记物-AGGAAAATGAGGTCTTCAATCTCAG，反向探针2：荧光标记物-CCTCAGAATGAGTGCTGACCGT；乙流病毒检测试剂系统的引物探针包括：正向外围引物：ACTTACCAATGGGTGCTTAA，反向外围引物：CGAAAAACAGAAAGGCAACAA，正向交叉引物：GCATCCCATTGGAACATGTCTTCAAATTTAGTAACATTGAAGGCTCAG，反向加速引物：CCTCAGAAGATGGCTGGTCAGTTTTCATAACCTCTTGGTCTC，反向探针1：荧光标记物-GTCTTCTTTTCCCAAAAGAAACTG，反向探针2：荧光标记物-CAAGAGCAGTGCTCAAACAAATGA；步骤二，采集测试者咽拭子样本；步骤三，将拭子头部浸入采样液中混匀；步骤四，将装置放置于水平台面，将采样液全部加入加样孔中，盖上加样孔盖；步骤五，反应温度50-65℃，静置10-30分钟；步骤六，随即将清洗液加入加样孔中，盖上加样孔盖，静置5-15分钟；步骤七，读取结果。
本发明的有益之处在于：
本发明的装置采用微流控导流设计，毛细管导流渠相互交叉，交叉的结构能够起到液体储存和缓冲的作用，因此可以实现各个流向的液体平衡；本发明的毛细管导流片的结构和材质的配合使得附着力大于内聚力，液体可以浸润接触面，并形成毛细管作用，从而充满两片毛细管导流片之间的空间，起到导流的作用；从而实现了液体流动的精准控制，提高精确性的同时避免了交叉污染，实现了核酸检测的居家自检；
本发明的装置无需外部电源、水源；无需专业实验室环境；无需专业培训；在不当操作的情况下亦不会发生触电、机械性伤害等危害性后果，操作简便适合普通群众进行居家式检测，以及一些野外环境或者缺乏专业人员、设备的条件下的检测；
本发明的装置在微流控，多重检测的支持下，能够进行定量检测；
本发明的装置具有高度的灵活性，可应用于传染病检测、病原体分型检测、环境微生物检测、生化指标检测、标志物检测、毒品检测等多个领域。
附图说明
图1是本发明上盖的一种实施例的俯视图；
图2是本发明上盖的一种实施例的仰视图；
图3是本发明上盖的一种实施例的侧视图；
图4是本发明夹层的一种实施例的上视图；
图5是本发明装置的一种实施例的截面图；
图6是本发明通断连接器的一种实施例的结构示意图；
图7是本发明通断连接器的另一种实施例的断开状态的结构示意图；
图8是本发明通断连接器的另一种实施例的连接状态的结构示意图；
图9是本发明的毛细管导流片的一种实施例的结构示意图；
图10是本发明的毛细管导流片的另一种实施例的结构示意图；
图中附图标记的含义：
1上盖，2底板，3夹层，4加样孔，5毛细管导流渠，51毛细管导流渠单元，511毛细管导流片，5111桥接部，512导流通道，501一级毛细管导流渠，502二级毛细管导流渠，503三级毛细管导流渠，6通断连接器，601第一缓冲槽，6011缺口，602第二缓冲槽，603隔断，604吸水垫，605连接槽，7加样槽，8功能腔室，801一级功能腔室，802二级功能腔室，9层析试纸，10加热模块，11结果阅读窗口，12电池，13开关，14功能腔室上盖，15开关孔，16指示灯窗口，17指示灯孔，18卡扣，19加样孔盖，20、21结果显示件固定组件。
具体实施方式
以下结合附图和具体实施例对本发明作具体的介绍。
如图1、2、3、4、5所示，一种自驱动微流控检测装置，包括：上盖1，固定于上盖1下的底板2，设置于上盖1和底板2之间的夹层3，设置于上盖1上的加样孔4，连接于加样孔4的微流控导流组件，设置于夹层3上并位置对应于微流控导流组件的加样槽7，设置于夹层3上并连接于微流控导流组件的功能腔室8，显示检测结果的结果显示件，设置于上盖1上并位于结果显示件上方的结果阅读窗口11，贴合于功能腔室8底部并设置于底板2内的加热模块10，连接于加热模块10的温控装置，连接于加热模块10并固定于底板2内的电源，连接于电源并固定于底板2内的开关13、指示灯，设置于上盖1上的开关孔15，设置于上盖1上的指示灯窗口16，设置于夹层3上的指示灯孔17，设置于上盖1、底板2上并用于组装的卡扣18，设置于上盖1、夹层3上的结果显示件固定组件20、21，设置于上盖1上并匹配于功能腔室8的功能腔室上盖14，固定于加样孔4上的加样孔盖19。结果显示件可以根据检测类型连接不同的零部件，可以连接上盖、夹层、底板等不受限制。
反应试剂：为检测时所用的物料，预置于功能腔室内，根据不同的检测项目，物料所含的物质和预装的方式可为多种组合。
结果显示件包括：层析试纸9、荧光检测仪器或试纸，可见染料检测试纸或装置，电信号检测仪等，无论是检测的方法还是检测的设备都不被限制等，只要能够连接反应后试剂显示出检测结果的产品都适用于本发明；其中层析试纸9用来呈现检测结果，以固定有检测线和质控线的条状纤维层析材料为固定相，测试液为流动相，荧光标记抗体或抗原固定于连接垫，通过毛细管作用使待分析物在层析条上移动和捕获从而进行检测。
功能腔室为给予发生物理反应或化学反应空间的腔室；不受检测类型和对象的限制。
加样孔4和加样孔盖19位于上盖1顶部的一端的外表面，加样孔4的内径和加样孔盖19下方柱形部分的外径匹配，通过盖上和打开加样孔盖19可实现加样孔4的开放和密闭。
功能腔室上盖14位于上盖1顶部的内表面，其形状和位置与夹层3上的功能腔室匹配。当装置完整组装后，功能腔室上盖14即可盖于功能腔室的上方，起到防止液体无序溢出的作用。
结果显示件固定组件20、21位于上盖1顶部的内表面和夹层3的上表面，由结果显示件固定凸起组成。当装置完整组装后，结果显示件固定凸起起到固定和托起免疫试纸的作用。
结果阅读窗口11位于上盖1的顶部，为透明材质，当装置完成组装时，结果阅读窗口11位于结果显示件的上方。当使用装置进行检测时，可通过结果阅读窗口11进行测试结果的读取。结果阅读窗口11可根据需要印刷引导结果阅读的图文。
上盖1和底板2上都设置有卡扣18，上盖1的卡扣18位于上盖1的侧部的内表面，用于和夹层3进行组装；底板2的卡扣18，用于和上盖1进行固定组装。
加样槽7位于夹层3的上表面，加样槽7包括：设置于的边缘位置的加样凸起，起到容纳液体的作用，形成于加样凸起之间并数条相互交叉的加样凹槽。当装置完整组装时，加样槽7与加样孔4位置对应，加样槽7的加样凹槽与上盖1的微流控导流组件的位置对应。
功能腔室一端与加样槽7相连，另一端与微流控导流组件相连。功能腔室为试剂与样本发生反应或混合的位置。
微流控导流组件连接加样孔4、功能腔室、结果显示件。微流控导流组件包括：毛细管导流渠5，连接于毛细管导流渠5并控制毛细管导流渠5的联通和断开的通断连接器6。
毛细管导流渠5由多个毛细管导流渠单元51组成；毛细管导流渠单元51包括：两片平行的毛细管导流片511，形成于毛细管导流片511之间的导流通道512。在导流通道512内的液体分子之间的吸引力为内聚力，在导流通道512内的液体分子与毛细管导流片511之间的吸引力为附着力；附着力大于内聚力。毛细管导流片511的材质和表面纹理不受限制，可以根 据检测的对象进行调节；作为一种实施例，毛细管导流片511的材质可为塑料，金属，玻璃，高分子聚合物等；毛细管导流片511的表面纹理可为光滑表面、磨砂表面及特定纹路。作为一种实施例，两片平行的毛细管导流片511之间的间距范围为：0.01-2mm。需要说明的是：这里的举例并非穷举，只要是能够让附着力大于内聚力的材质、纹理、间距都在本发明的保护范围内。
如图9、10所示，毛细管导流片511的端部设置有向下延伸的桥接部5111，作为一种实施例，如图9所示，毛细管导流片511的一端设置有一个桥接部5111，桥接部5111向下弯曲，桥接部5111使得毛细管导流片511的末端与功能腔室中预装的反应物料接触。作为一种实施例，如图10所示，毛细管导流片511的两端设置有两个桥接部5111，桥接部5111向下延伸，整体呈U型。
毛细管导流渠5在加样孔4下方位置相互交叉设置，交叉点位于加样孔4中心的下方。
该装置亦可根据不同的检测需求，功能腔室和毛细管导流渠的构成可为多级结构：如图6、7所示，具体的结构体现如下所述。
作为一种实施例，如图6所示，毛细管导流渠5为三级毛细管导流渠503，功能腔室8为二级功能腔室802；三级毛细管导流渠503分别是：连接于加样槽7与一级功能腔室801之间的一级毛细管导流渠501，连接于通断连接器6与二级功能腔室802之间的二级毛细管导流渠502，连接于二级功能腔室802与层析试纸9之间的三级毛细管导流渠503。如图6所示，通断连接器6包括：连接于一级功能腔室801的第一缓冲槽601，连接于第一缓冲槽601并放置二级毛细管导流渠502的第二缓冲槽602，设置于第一缓冲槽601与第二缓冲槽602之间的隔断603；隔断603的高度低于第一缓冲槽601、第二缓冲槽602的高度。第一缓冲槽601与一级功能腔室801之间设置有缺口6011，引导液体流向第一缓冲槽601。运行过程为：当测试样本从加样孔4加入后，经一级毛细管导流渠501引导至一级功能腔室801进行反应；反应结束后，通过加入更多的液体，使得液体从一级功能腔室801溢出进入第一缓冲槽601，当一级功能腔室801中加入的样本略微过量时，液体从一级功能腔室801的缺口6011溢出至第一缓冲槽601，此时第一缓冲槽601起到隔断603一级功能腔室801与第二缓冲槽602的作用，因为隔断603的高度低于第一缓冲槽601、第二缓冲槽602的高度，当后续继续加入更多的液体时，进入第一缓冲槽601的液体越过隔断603进入第二缓冲槽602，从而实现通断连接器6的连通，经由二级毛细管导流渠502进入二级功能腔室802进行反应，此后通过三级毛细管导流渠503将液体引导至层析试纸9获得结果。
作为另一种实施例，通断连接器6包括：连接于一级功能腔室801的连接槽605，放置在连接槽605内并对应于二级毛细管导流渠502下的吸水膨胀件。作为一种实施例，吸水膨胀件为吸水垫604；需要说明是：吸水膨胀件的材质形状不受限制，只要是能够吸水膨胀实现通断都可以应用于本发明。检测肛拭子中肠炎沙门氏菌时，如图7所示，断开状态时，样本加入后均匀分配至一级功能腔室801中进行反应，一级功能腔室801内的液体通过缺口6011流入连接槽605，吸水垫604吸水膨胀，如图8所示，进入连接状态，吸水垫604接触二级毛细管导流渠502，通过二级毛细管导流渠502将两个反应的扩增产物导流至二级功能腔室802，在二级功能腔室802中进行混合，而后通过三级毛细管导流渠503，将二级功能腔室802与核酸免疫层析试纸9相连，层析试纸9的C线捕获GAPDH系统阳性扩增产物，T线捕获肠炎沙门氏菌扩增系统阳性产物。需要说明的是：这里的一级、二级、三级仅是为了清楚说明结构，并非必须是具有二级功能腔室和三级毛细管导流渠才能使用如上所述的通断连接器，功能腔室和毛细管导流渠的个数是不受限制的。
需要说明的是本发明只举了两个例子，但是这都不是穷举，只要是能够实现试剂液体与毛细管导流渠之间的连接和断开的都受到本发明的启示，都在本发明的保护范围内。除了本发明这样的结构，还可以是在二级毛细管导流渠502与一级功能腔室801之间设置连接管，连接管倾斜或垂直连接于一级功能腔室801，在液体流量达到指定的量后才能连通二级毛细管导流渠502，以此实现通断。也可以利用液压实现通断。也可以通过机械装置实现通断，比如阀门等；膨胀件也可以通过遇热膨胀的原理进行膨胀，在此不一一举例。
加热模块10组件包括：贴合于功能腔室底部并设置于底板2内的加热模块10，连接于加热模块10的温控装置，连接于加热模块10并固定于底板2内的电源，连接于电源并固定于底板2内的开关13、指示灯，设置于上盖1上的开关孔15，设置于上盖1上的指示灯窗口16，设置于夹层3上的指示灯孔17。
电池12，加热模块10、指示灯和开关13固定于底板2的内部。加热模块10：起到为反应提供加热的功能，它与功能腔室的底部贴合，通过电加热和温控装置控制功能腔室的温度。电池12：电池12预装于夹层3和底板2之间的空间，通过电线连接加热模块10、开关13和指示灯，为加热模块10提供电源。指示灯：指示灯起到指示电源是否接通的作用，当电源接通时，指示灯亮起。指示灯窗口16位于上盖1加样孔4一端，为透光材质，其在上盖1内表面的部分具有凸起结构，用于固定指示灯。开关孔15位于上盖1的侧部加样孔4一端，用于安装开关13。指示灯孔17为容纳指示灯的圆孔位于装置的加样孔4一端；开关13：为装置的电源开关13，通过开关13的打开和关闭可以接通和切断装置的电流，从而控制检测反应的进行。
该装置可根据不同的检测需求调整通道数量，如：2通道自驱动微流控检测装置为加样孔4通过毛细管导流渠5与两个功能腔室连接，每个功能腔室对应一条层析试纸9，一个结果阅读窗。4通道自驱动微流控检测装置为加样孔4通过毛细管导流渠5与四个功能腔室连接，每个功能腔室对应一条层析试纸9，一个结果阅读窗。6通道自驱动微流控检测装置为加样孔4通过毛细管导流渠5与6个功能腔室相连，每个功能腔室对应一条层析试纸9，一个结果阅读窗。
需要说明的是：装置可以根据检测项目的需求，选择使用各个组件。如当检测反应不需要加热时可不使用电池12，加热模块10，开关13等以节省制造成本。
本发明装置的操作步骤包括：
1.加样
将检测装置至于水平台面上，打开加样孔盖19，将样本加入加样孔4中。液体在微流控导流装置的引导下，均匀流向各个功能腔室。
2.检测反应
盖上加样孔盖19，打开开关13(如采用非加热检测方法则不需开关13)，静置0-60分钟。
3.层析反应
检测反应完成后，随即打开加样孔盖19，加入缓冲液，使得装置内部的液体从功能腔室流动至层析试纸9。重新盖上上盖1，静置5-15分钟。
4.从样本读取窗口读取测试结果。
以下通过本装置的几个检测应用验证本装置的检测精确度。
应用一，一种自驱动微流控核酸检测装置应用于新冠/新冠突变株/甲流/乙流病毒核酸检测的方法。
检测项目：新型冠状病毒、新型冠状病毒突变株B.1.1.7、甲流病毒、乙流病毒。
1，材料和方法
采用4通道自驱动微流控检测装置。4通道自驱动微流控检测装置为加样孔4通过毛细管导流渠5与四个功能腔室连接，每个功能腔室对应一条层析试纸9，一个结果阅读窗。
根据检测液体的特性，该方案中微流控导流装置的材质为PE材质，导流片的间距为0.2mm。
采用的层析试纸9为核酸免疫层析试纸9。
引物和探针如下表1所示：
表1
Figure PCTCN2021093862-appb-000001
Figure PCTCN2021093862-appb-000002
DNA聚合酶：Bst 3.0DNA聚合酶(NEB)
反转录酶：AMV反转录酶(NEB)
每个功能腔室中预装的成分为：正向外围引物：1-50pmol，反向外围引物：1-50pmol，正向交叉引物：10-500pmol，反向交叉引物：10-500pmol，正向探针：1-100pmol，反向探针：1-100pmol，DNA聚合酶:5-50U，反转录酶:0.5-8U，dNTP:10-300nmol，BSA:0.1-5μg。
四个检测系统的物料，按照系统的成份组成，依次预装于4个功能腔室中。
采样液成分：5-100mM Tris-HCl，0.5-40μM DTT，0.1～1M Betaine，0.1％～5％TritonX-100，2～15mM MgSO 4，pH 7-9@25℃。清洗液成分：5-100mM Tris-HCl，2～15mM NaCl。
检测方法：
1，采集测试者咽拭子样本；
2，将拭子头部浸入0.2-1ml的采样液中，转动混匀或摇动混匀5-15秒；
3，将装置放置于水平台面，将采样液全部加入加样孔4中，盖上加样孔盖19；
4，打开电源开关13，反应温度50-65℃，静置10-30分钟；
5，步骤4完成后，随即将清洗液0.2-1ml加入加样孔4中，盖上加样孔盖19，静置5-15分钟；
6，读取结果。
结果：
使用分别含有新型冠状病毒，新型冠状病毒突变型B.1.1.7,甲流病毒，乙流病毒四种病原体核酸的假病毒作为检测样本进行检测；在采集的拭子上加入假病毒，样本浓度为100000拷贝/每拭子，10000拷贝/拭子，1000拷贝/拭子，100拷贝/拭子，10拷贝/拭子，1拷贝/拭子。每种假病毒，每个样本浓度的重复数为10(n＝10)。检测设阴性对照10测试。
检测结果显示：
当样本浓度为100000拷贝/拭子时，新型冠状病毒假病毒，新型冠状病毒B.1.1.7假病毒,甲流病毒假病毒，乙流病毒假病毒全部检出，检出率均为100％。
当样本浓度为10000拷贝/拭子时，新型冠状病毒假病毒，新型冠状病毒B.1.1.7假病毒,甲流病毒假病毒，乙流病毒假病毒全部检出，检出率均为100％。
当样本浓度为1000拷贝/拭子时，新型冠状病毒假病毒，新型冠状病毒B.1.1.7假病毒,甲流病毒假病毒，乙流病毒假病毒全部检出，检出率均为100％。
当样本浓度为100拷贝/拭子时，新型冠状病毒假病毒检出率为80％；新型冠状病毒B.1.1.7假病毒检出率为50％；甲流病毒假病毒检出率为30％；乙流病毒假病毒检出率为40％；
当样本浓度为10拷贝/拭子时，新型冠状病毒假病毒，新型冠状病毒B.1.1.7假病毒,甲流病毒假病毒，乙流病毒假病毒全部未检出，检出率均为0％。
当样本浓度为1拷贝/拭子时，新型冠状病毒假病毒，新型冠状病毒B.1.1.7假病毒,甲流病毒假病毒，乙流病毒假病毒全部未检出，检出率均为0％。
阴性对照结果为阴性。
结论：
实验结果证明一种自驱动微流控检测装置，应用于新型冠状病毒，新型冠状病毒突变型B.1.1.7,甲流病毒，乙流病毒四种核酸检测，灵敏度达到或优于1000拷贝/拭子。
应用二，一种自驱动微流控核酸检测装置应用于检测食品微生物的方法。
检测项目：大肠杆菌、肠炎沙门氏菌。
1.材料和方法
采用2通道自驱动微流控检测装置，2通道自驱动微流控检测装置为加样孔4通过毛细管导流渠5与两个功能腔室连接，
每个功能腔室对应一条层析试纸9，一个结果阅读窗。
根据检测液体的特性，该方案的微流控导流装置为玻璃材质，导流片之间的间距为1mm。
采用的层析试纸9为核酸免疫层析试纸9。
引物和探针如下表2所示：
表2
Figure PCTCN2021093862-appb-000003
DNA聚合酶：Bst 3.0DNA聚合酶(NEB)
每个功能腔室中预装的成分为：
正向外围引物：1-50pmol，反向外围引物：1-50pmol，正向交叉引物：10-500pmol，反向交叉引物：10-500pmol，正向探针：1-100pmol，反向探针：1-100pmol，DNA聚合酶:5-50U，dNTP:10-300nmol，BSA:0.1-5μg。
两个检测系统的物料，按照系统的成份组成，依次预装于两个功能腔室中。
采样液成分：5-100mM Tris-HCl，0.5-40μM DTT，0.1～50mg/ml BSA，0.1～1M Betaine，0.1％～5％TritonX-100，2～15mM MgSO 4，pH 7-9@25℃。清洗液成分：5-100mM Tris-HCl，2～15mM NaCl。
检测方法：
步骤一，使用沾有少量采样液的拭子对食材表面进行大范围擦拭5-20次；
步骤二，将拭子头部浸入0.2-1ml的采样液中，转动混匀或摇动混匀5-15秒；
步骤三，将装置放置于水平台面，将采样液全部加入加样孔4中，盖上加样孔盖19；
步骤四，打开电源开关13，反应温度50-75℃，静置10-30分钟；
步骤五，步骤四完成后，随即将清洗液0.2-1ml加入加样孔4中，盖上加样孔盖19，静置5-15分钟；
步骤六，读取结果。
结果：
使用3批被大肠杆菌污染的食材、3批被肠炎沙门氏菌污染的食材、和三批没有被大肠杆菌和肠炎沙门氏菌污染的食材。采集的拭子样本一式两份，一份按照上述操作方法采集样本，并进行检测。另一份使用qPCR方法进行细菌含量的测定，三批被大肠杆菌污染的食材上采集的拭子标本中菌含量为：13500菌/拭子、6500菌/拭子、16000菌/拭子；三批被肠炎沙门氏菌污染的食材上采集的拭子标本中菌含量为：3500菌/拭子、1200菌/拭子、700菌/拭子。
自驱动微流控核酸检测装置应用于检测食品微生物的方法进行检测的结果如下表3：
表3
Figure PCTCN2021093862-appb-000004
结论：
检测结果显示：自驱动微流控检测装置应用于检测食品微生物的方法能够一小时内检测食材是否遭到特定微生物的污染，结果准确，其检测灵敏度优于1000菌/拭子。
应用三，一种自驱动微流控检测装置应用于检测尿液样本中微生物的方法
检测项目：解脲支原体、金黄色葡萄球菌、淋病奈瑟球菌、白色念珠菌
2，材料和方法
采用4通道自驱动微流控核酸检测装置，4通道自驱动微流控检测装置为加样孔4通过毛细管导流渠5与四个功能腔室连接，每个功能腔室对应一条层析试纸9，一个结果阅读窗。
根据检测液体的特性，该方案采用的微流控导流装置为PE材质，导流片之间的间距为0.5mm。
采用的层析试纸9为核酸免疫层析试纸9。
引物探针如下表4所示：
表4
Figure PCTCN2021093862-appb-000005
DNA聚合酶：Bst 3.0DNA聚合酶(NEB)
每个功能腔室中预装的成分为：正向外围引物：1-50pmol，反向外围引物：1-50pmol，正向交叉引物：10-500pmol，反向交叉引物：10-500pmol，正向探针：1-100pmol，反向探针：1-100pmol，DNA聚合酶:5-50U，dNTP:10-300nmol，BSA:0.1-5μg。
四个检测系统的物料，按照系统的成份组成，依次预装于4个功能腔室中。
采样液成分：5-100mM Tris-HCl，0.5-40μM DTT，0.1～50mg/ml BSA，0.1～1M Betaine，0.1％～5％TritonX-100，2～15mM MgSO 4，pH 7-10@25℃。清洗液成分：5-100mM Tris-HCl，2～15mM NaCl。
检测方法：
步骤一，使用干燥的拭子头部沾取尿液样本，当拭子顶部吸收到样本即可取出；
步骤二，将拭子头部浸入0.2-1ml的采样液中，转动混匀或摇动混匀5-15秒后静置5-10分钟；
步骤三，将装置放置于水平台面，将采样液全部加入加样孔4中，盖上加样孔盖19；
步骤四，打开电源开关13，静置30分钟；
步骤五，步骤四完成后，随即将清洗液0.2-1ml加入加样孔4中，盖上加样孔盖19，静置5-15分钟；
步骤六，读取结果。
结果：
对含有解脲支原体、金黄色葡萄球菌、淋病奈瑟球菌、白色念珠菌的尿液进行检测，尿液中各病原体经稀释调整为100000cfu/ml、10000cfu/ml、5000cfu/ml、2000cfu/ml、1000cfu/ml、500cfu/ml、200cfu/ml。每种病原体，每个浓度的重复数为10(n＝10)。
当样本浓度为100000cfu/ml时，解脲支原体、金黄色葡萄球菌、淋病奈瑟球菌、白色念珠菌，检出率均为100％。
当样本浓度为10000cfu/ml时，解脲支原体、金黄色葡萄球菌、淋病奈瑟球菌、白色念珠菌，检出率均为100％。
当样本浓度为5000cfu/ml时，解脲支原体、金黄色葡萄球菌、淋病奈瑟球菌、白色念珠菌，检出率均为100％。
当样本浓度为2500cfu/ml时，解脲支原体、金黄色葡萄球菌、淋病奈瑟球菌检出率为100％，白色念珠菌检出率为60％。
当样本浓度为1000cfu/ml时，解脲支原体检出率为100％，金色葡萄球菌检出率为80％，淋病奈瑟球菌检出率为50％。白色念珠菌检出率为30％。
当样本浓度为500cfu/ml时，解脲支原体检出率为50％，金色葡萄球菌检出率为40％，淋病奈瑟球菌检出率为20％，白色念珠菌未检出。
当样本浓度为250cfu/ml时，解脲支原体检出率为10％，金色葡萄球菌、淋病奈瑟球菌、白色念珠菌均未检出。
结论：
实验结论证明，自驱动微流控检测装置应用于检测尿液样本中微生物的方法可以检出尿液中的病原体。其检出下限分别为：解脲支原体250cfu/ml，金色葡萄球菌500cfu/ml，淋病奈瑟球菌500cfu/ml，白色念珠菌1000cfu/ml。
应用四，一种自驱动微流控检测装置应用于定量检测肾脏标志物的方法。
检测项目：视黄醇结合蛋白(RBP)
材料和方法：
采用4通道自驱动微流控检测装置，4通道自驱动微流控检测装置为加样孔4通过毛细管导流渠5与四个功能腔室连接，每个功能腔室对应一条层析试纸9，一个结果阅读窗。
根据检测液体的特性，该方案中采用的微流控导流装置的导流片为PP材质，导流片的间距为0.4mm。
试剂和制备的步骤：
步骤一，将0.01-0.1％氯酸金溶液加热至沸腾后，在搅拌状态下加入1％柠檬酸三钠，二者体积比为10/1-100/1；当溶液完全变为透明的红色时，继续煮沸5-30分钟，搅拌冷却后，得到所需的纳米金。
步骤二，将步骤1中得到的纳米金溶液用碳酸钾溶液调节pH值至8-10，将鼠抗RBP单克隆抗体1，加入溶液均匀混合，浓度为20-50ng/μl.室温搅拌5-40分钟后，加10％入小牛血清溶液，其体积比为10/1-100/1搅拌20分钟。
步骤三，将步骤2制得的溶液以10000-17000rpm/min、低温离心10-30分钟、去上清。沉淀使用含5％BSA的硼酸盐缓冲液重悬至原体积，再次以同样的方法离心去上清后，加含5％BSA的硼酸盐缓冲液重悬至原体积的1/20，得到金标RBP单克隆抗体1。
步骤四，将NC膜附在PVC底板2，用往复划膜仪在NC膜的不同位置喷点RBP单克隆堂提2及羊抗鼠二抗，分别为检测线(T线)和质控线(C线)，37℃烘干2小时。
步骤五，在步骤3得到的金标RBP单克隆抗体溶液中加入PVP,PEG,蔗糖0.02g/mL,0.02g/mL,0.5g/mL。然后喷于玻璃纤维膜的结合垫上，37℃烘干备用。
步骤六，玻璃纤维素膜的样品垫、结合垫、NC膜和吸水垫604组装成为检测试纸条。通过调整结合垫和测试线的抗体浓度，将该试纸条的灵敏度下限调整为100mg/L。
步骤七，将检测试纸条组装于自驱动微流控检测装置中。
步骤八，在4个功能腔室中依次预装0.2475mL、0.225mL、0.125mL、0mL的生理盐水。
步骤九，将0.6mL的尿液样本加入检测装置的加样孔4中，1分钟后将孔中剩余的样本吸出。在微流控导流装置的控制下，进入4个功能腔室中，并且每个腔室的最终体积为0.25mL。因此样本在4个功能腔室中分别被稀释100倍，10倍稀释，2倍稀释，和不稀释。
步骤十，读取结果。
结果：分别测试4个样本，其中RBP的含量分来自健康人群，A样本浓度为150mg/L,B样本300mg/L，和有肾脏炎症的人群，RBP浓度为C样本1200mg/L和D样本1600mg/L.
检测装置的结果如表5所示：
表5
样本 1号试纸 2号试纸 3号试纸 4号试纸
A - - - +
B - - + +
C - + + +
D - + + +
结果显示A样本的RBP浓度大于100mg/L,B样本浓度大于200mg/L，C样本的浓度大于1000mg/L，D样本的浓度大于1000mg/L。实验结果提示两位患有肾脏验证的受试者，肾脏存在炎症，与实际样本相符。
结论：本发明自驱动微流控检测装置可以用于检测肾脏标志物，并对其进行初步的定量，为肾脏的健康提供初步的判断依据。
应用五，一种自驱动微流控核酸检测装置应用于甲流和乙流检测的方法。
检测项目：甲流病毒、乙流病毒。
1，材料和方法
采用2通道多级自驱动微流控检测装置，2通道自驱动微流控检测装置为加样孔4通过毛细管导流渠5与两个功能腔室连接，每个功能腔室对应一条层析试纸9，一个结果阅读窗。
样本从加样孔4进入后，经一级微流控导流渠进入一级功能腔室801，在一级功能腔室801中由加热装置进行加热进行核酸扩增反应，再通过加入清洗液，使得加入液体的体积超过通断连接器6的灵界值，从而启动通断连接器6使得液体经由二级微流控导流渠进入二级功能腔室802，在二级功能腔室802中，两种反应物充分混合，再经由三级微流控导流渠流至核酸免疫层析试纸9。在核酸免疫试纸上设置有两条T线，分别显示甲流和乙流的检测结果。
根据采样液的特性该方案中，微流控导流装置采用玻璃材质，导流片之间的间距为0.2mm。
采用的层析试纸9为核酸免疫层析试纸9，甲乙流的检测结果在一条试纸条上呈现。
引物和探针如下表6所示：
表6
Figure PCTCN2021093862-appb-000006
DNA聚合酶：Bst 3.0DNA聚合酶(NEB)；反转录酶：AMV反转录酶(NEB)；每个功能腔室中预装的成分为：正向外围引物：1-50pmol，反向外围引物：1-50pmol，正向交叉引物：10-500pmol，反向交叉引物：10-500pmol，正向探针：1-100pmol，反向探针：1-100pmol；DNA聚合酶:5-50U；反转录酶:0.5-8U；dNTP:10-300nmol；BSA:0.1-5μg。
两个检测系统的物料，按照系统的成份组成，依次预装于两个功能腔室中。
采样液成分：5-100mM Tris-HCl，0.5-40μM DTT，0.1～1M Betaine，0.1％～5％TritonX-100，2～15mM MgSO 4，pH 7-9@25℃。清洗液成分：5-100mM Tris-HCl，2～15mM NaCl。
检测方法：
1，采集测试者咽拭子样本；
2，将拭子头部浸入0.2-1ml的采样液中，转动混匀或摇动混匀5-15秒；
3，将装置放置于水平台面，将采样液全部加入加样孔4中，盖上加样孔盖19；
4，打开电源开关13，反应温度50-65℃，静置10-30分钟；
5，步骤4完成后，随即将清洗液0.2-1ml加入加样孔4中，盖上加样孔盖19，静置5-15分钟；
6，读取结果。
结果：
使用分别含有甲流和乙流病毒四的假病毒作为检测样本进行检测；在采集的拭子上加入假病毒，样本浓度为100000拷贝/每拭子，10000拷贝/拭子，1000拷贝/拭子，100拷贝/拭子，10拷贝/拭子。每种假病毒，每个样本浓度的重复数为10(n＝20)。检测设阴性对照10测试。
检测结果显示：
当样本浓度为100000拷贝/拭子时，甲流病毒假病毒，乙流病毒假病毒全部检出，检出率均为100％。
当样本浓度为10000拷贝/拭子时，甲流病毒假病毒，乙流病毒假病毒全部检出，检出率均为100％。
当样本浓度为1000拷贝/拭子时，甲流病毒假病毒检出率为85％，乙流病毒假病毒检出率均为90％。
当样本浓度为100拷贝/拭子时，甲流病毒假病毒检出率为30％；乙流病毒假病毒检出率为40％；
当样本浓度为10拷贝/拭子时，甲流病毒假病毒，乙流病毒假病毒全部未检出，检出率均为0％。
阴性对照结果为阴性。
结论：
实验结果证明一种自驱动微流控检测装置，能够应用于甲流病毒，乙流病毒四种核酸检测，且结果通过多级的微流控导流装置最终呈现在一条试纸条上，结果清晰明确，更加直观。
应用六，自驱动微流控核酸检测装置用于检测肛拭子中肠炎沙门氏菌的方法。
检测项目：肠炎沙门氏菌
1材料和方法
采用2通道自驱动微流控检测装置。
根据检测液体的特性，该方案的微流控导流装置为玻璃材质，导流片之间的间距为0.5mm，采用的开关13装置为吸水垫604开关13装置。
如7、8所示，当加入液体较少时，开关13装置中的吸水垫604，因为吸收的液体较少，高度较低于二级毛细管导流渠502，无法连接。当操作中加入更多的液体时，吸水垫604的高度增加，并与二级毛细管导流渠502相连，从而连通二级毛细管导流渠502，将液体输送至二级功能腔室802，在二级功能腔室802中进行混合。而后通过三级毛细管导流渠503，将二级功能腔室802与核酸免疫层析试纸9相连，层析试纸9的C线捕获GAPDH系统阳性扩增产物，T线捕获肠炎沙门氏菌扩增系统阳性产物。
采用的层析试纸9为核酸免疫层析试纸9。
引物和探针如下表7所示：
表7
Figure PCTCN2021093862-appb-000007
DNA聚合酶：Bst 3.0DNA聚合酶(NEB)
每个功能腔室中预装的成分为：
正向外围引物：1-50pmol；反向外围引物：1-50pmol；正向交叉引物：10-500pmol；反向交叉引物：10-500pmol；正向探针：1-100pmol；反向探针：1-100pmol；DNA聚合酶:5-50U；dNTP:10-300nmol；BSA:0.1-5μg；两个检测系统的物料，按照上述成分组成，依次预装于2个功能腔室中。
采样液成分：
5-100mM Tris-HCl，0.5-40μM DTT，0.1～50mg/ml BSA，0.1～1M Betaine，0.1％～5％TritonX-100，2～15mM MgSO 4，pH 7-9@25℃；清洗液成分：5-100mM Tris-HCl，2～15mM NaCl。
检测方法：
步骤一，采集肛拭子；
步骤二，将拭子头部浸入0.2-1ml的采样液中，转动混匀或摇动混匀5-15秒，使用滤膜对采样液进行过滤，滤去大体积的残渣；
步骤三，将装置放置于水平台面，将采样液全部加入加样孔4中，盖上加样孔盖19；
步骤四，打开电源开关13，反应温度50-75度，静置10-30分钟；
步骤五，步骤四完成后，随即将清洗液0.2-1ml加入加样孔4中，盖上加样孔盖19，静置5-15分钟；
步骤六，读取结果。
结果：
上述操作方法采集样本，并进行检测。
自驱动微流控核酸检测装置应用于检测肛拭子中肠炎沙门氏菌的结果如下表8：
表8
Figure PCTCN2021093862-appb-000008
结论：
检测结果显示：自驱动微流控检测装置应用于检测肛拭子中肠炎沙门氏菌，能稳定检出样本浓度为1000菌/拭子的样本。证明该装置能够用于检测肛拭子中肠炎沙门氏菌。
本发明采用微流控导流设计，实现了液体流动的精准控制，结果呈现明确，报告时间短，无需专业培训，无需昂贵仪器亦可得到准确的检测结果。该产品在不当操作时，不会产生危害性的后果，具有极高的安全性，十分适合普通群众在家庭进行自我检测。该产品可对尿液，口咽拭子、肛拭子、食材表面刮取拭子等样本进行检测，具有十分广泛的适用范围。通过家庭自检，选择合适的应对措施，可缓解群众的焦虑，避免不必要的医疗资源占用和人群聚集。
以上显示和描述了本发明的基本原理、主要特征和优点。本行业的技术人员应该了解，上述实施例不以任何形式限制本发明，凡采用等同替换或等效变换的方式所获得的技术方案，均落在本发明的保护范围内。

Claims (19)

  1. 一种自驱动微流控检测装置，其特征在于，包括：上盖，固定于上盖下的底板，设置于上盖和底板之间的夹层，设置于上盖上的加样孔，连接于加样孔的微流控导流组件，设置于夹层上并位置对应于微流控导流组件的加样槽，设置于夹层上并连接于微流控导流组件的功能腔室，显示检测结果的结果显示件；所述微流控导流组件包括：毛细管导流渠，连接于毛细管导流渠之间并控制毛细管导流渠的联通和断开的通断连接器。
  2. 根据权利要求1所述的一种自驱动微流控检测装置，其特征在于，所述毛细管导流渠由多个毛细管导流渠单元组成；所述毛细管导流渠单元包括：两片平行的毛细管导流片，形成于毛细管导流片之间的导流通道。
  3. 根据权利要求2所述的一种自驱动微流控检测装置，其特征在于，所述两片平行的毛细管导流片之间的间距范围为：0.01-10mm。
  4. 根据权利要求2所述的一种自驱动微流控检测装置，其特征在于，在所述导流通道内的液体分子之间的吸引力为内聚力，在所述导流通道内的液体分子与毛细管导流片之间的吸引力为附着力；所述附着力大于内聚力。
  5. 根据权利要求2所述的一种自驱动微流控检测装置，其特征在于，所述毛细管导流片的端部设置有向下延伸的桥接部。
  6. 根据权利要求1所述的一种自驱动微流控检测装置，其特征在于，所述毛细管导流渠为三级毛细管导流渠，功能腔室为二级功能腔室；三级毛细管导流渠分别是：连接于加样槽与一级功能腔室之间的一级毛细管导流渠，连接于通断连接器与二级功能腔室之间的二级毛细管导流渠，连接于二级功能腔室与结果显示件之间的三级毛细管导流渠。
  7. 根据权利要求6所述的一种自驱动微流控检测装置，其特征在于，所述一级功能腔室设置有缺口。
  8. 根据权利要求1所述的一种自驱动微流控检测装置，其特征在于，所述通断连接器包括：连接于功能腔室的第一缓冲槽，连接于第一缓冲槽并放置毛细管导流渠的第二缓冲槽，设置于所述第一缓冲槽与第二缓冲槽之间的隔断；所述隔断的高度低于第一缓冲槽、第二缓冲槽的高度。
  9. 根据权利要求1所述的一种自驱动微流控检测装置，其特征在于，所述通断连接器包括：连接于功能腔室的连接槽，放置在连接槽内并对应于毛细管导流渠下的吸水膨胀件。
  10. 根据权利要求1所述的一种自驱动微流控检测装置，其特征在于，所述毛细管导流渠在加样孔下方位置相互交叉设置，交叉点位于加样孔中心的下方。
  11. 根据权利要求1所述的一种自驱动微流控检测装置，其特征在于，还包括：设置于上盖上 并位于结果显示件上方的结果阅读窗口，贴合于功能腔室底部并设置于底板内的加热模块，连接于加热模块的温控装置，连接于加热模块并固定于底板内的电源，连接于电源并固定于底板内的开关、指示灯，设置于上盖上的开关孔，设置于上盖上的指示灯窗口，设置于夹层上的指示灯孔。
  12. 根据权利要求1所述的一种自驱动微流控检测装置，其特征在于，还包括：设置于上盖、底板上并用于组装的卡扣，设置于上盖、夹层上的结果显示件固定组件，设置于上盖上并匹配于功能腔室的功能腔室上盖，固定于加样孔上的加样孔盖。
  13. 根据权利要求1所述的一种自驱动微流控检测装置的用途，其特征在于，装置预装有核酸检测试剂并用于生物核酸检测。
  14. 根据权利要求1所述的一种自驱动微流控检测装置的用途，其特征在于，用于核酸检测新冠/新冠突变株/甲流/乙流病毒核酸检测包括如下内容：
    采用4通道自驱动微流控检测装置，4通道自驱动微流控检测装置为加样孔通过毛细管导流渠与四个功能腔室连接，每个功能腔室对应一条层析试纸，一个结果阅读窗；
    所述毛细管导流渠的毛细管导流片的材质为PE材质，毛细管导流片的间距为0.2mm；
    采用的层析试纸为核酸免疫层析试纸；
    步骤一，将新冠检测试剂系统，新冠B.1.1.7检测试剂系统，甲流检测试剂系统，乙流检测试剂系统预装于4个功能腔室中；
    所述新冠检测试剂系统的引物探针包括：
    正向外围引物：GGCAGTCAAGCCTCTTCTC，
    反向外围引物：TCTGTCAAGCAGCAGCAAAG，
    正向交叉引物：TTCCCCTACTGCTGCCTGGAGTTCCTCATCACGTAGTCGC，
    反向加速引物：AAGAGCAGCATCACCG，
    反向探针1：荧光标记物-GAATTTCTTGAACTGTTGCG，
    反向探针2：荧光标记物-TTCCCCTACTGCTGCCTGGA；
    所述新冠B.1.1.7检测试剂系统的引物探针包括：
    正向外围引物：TTCTTTCACACGTGGTGT，
    反向外围引物：GACAGGGTTATCAAACCTCT，
    正向交叉引物：AGGTAAGAACAAGTCCTGAGTTGATTATTACCCTGACAAAGTTTTCAG，
    反向加速引物：AGTACCATTGGTCCCAGA，
    反向探针1：荧光标记物-GTCCCAGAGATAGCATGG，
    反向探针2：荧光标记物-AGGTAAGAACAAGTCCTGAGTTGA；
    所述甲流检测试剂系统的引物探针包括：
    正向外围引物：CAGAGGGCAATGATGGATCA，
    反向外围引物：ATCCCGACCAGTGAGTACC，
    正向交叉引物：CCTCAGAATGAGTGCTGACCGTAAGTCGAAACCCAGGAAACG，
    反向加速引物：CTTCCCTTTCAAAGTCATGCCCA，
    反向探针1：荧光标记物-AGGAAAATGAGGTCTTCAATCTCAG，
    反向探针2：荧光标记物-CCTCAGAATGAGTGCTGACCGT；
    所述乙流检测试剂系统的引物探针包括：
    正向外围引物：ACTTACCAATGGGTGCTTAA，
    反向外围引物：CGAAAAACAGAAAGGCAACAA，
    正向交叉引物：CATCCCATTGGAACATGTCTTCAAATTTAGTAACATTGAAGGCTCAG，
    反向加速引物：CCTCAGAAGATGGCTGGTCAGTTTTCATAACCTCTTGGTCTC，
    反向探针1：荧光标记物-GTCTTCTTTTCCCAAAAGAAACTG，
    反向探针2：荧光标记物-CAAGAGCAGTGCTCAAACAAATGA；
    步骤二，采集测试者咽拭子样本；
    步骤三，将拭子头部浸入采样液中混匀；
    步骤四，将装置放置于水平台面，将采样液全部加入加样孔中，盖上加样孔盖；
    步骤五，反应温度50-65℃，静置10-30分钟；
    步骤六，随即将清洗液加入加样孔中，盖上加样孔盖，静置5-15分钟；
    步骤七，读取结果。
  15. 根据权利要求1所述的一种自驱动微流控检测装置的用途，其特征在于，用于大肠杆菌、肠炎沙门氏菌检测包括如下内容：
    采用2通道自驱动微流控检测装置，2通道自驱动微流控检测装置为加样孔通过毛细管导流渠与两个功能腔室连接，每个功能腔室对应一条层析试纸，一个结果阅读窗；
    所述毛细管导流渠的毛细管导流片的材质为玻璃材质，毛细管导流片之间的间距为1mm；
    采用的层析试纸为核酸免疫层析试纸；
    步骤一，将大肠杆菌检测试剂系统，肠炎沙门氏菌检测试剂系统预装于2个功能腔室中；
    所述大肠杆菌检测试剂系统的引物探针包括：
    正向外围引物：ACCGTCAGGAAGCGGTAC，
    反向外围引物：TTTCACCCACTCTTCCTGGAT，
    正向交叉引物：AGACGGTTGGAGTTGGAGGAGTGCAGAACAGGCGGAAGTT，
    反向加速引物：GTCTTTCGCATCGTCAATCAAAA，
    反向探针1：荧光标记物-TTTTCGAACCGACCACCAACAC，
    反向探针2：荧光标记物-AGACGGTTGGAGTTGGAGGAGT；
    所述肠沙门氏菌检测试剂系统的引物探针包括：
    正向外围引物：CGTGATGCTGAAAGTACCGA，
    反向外围引物：GGCCGCCAAAACTTTCCTGA，
    正向交叉引物：CCACCGCGTACGGACTTCACCGAAACACAAACGGGCAAG，
    反向加速引物：AGATCTTTTAGCAATTGCTTCT，
    反向探针1：荧光标记物-TGCCGCGCATACGGAACAG，
    反向探针2：荧光标记物-CCACCGCGTACGGACTTCAC；
    步骤二，使用沾有少量采样液的拭子对待测物表面进行大范围擦拭5-20次；
    步骤三，将拭子头部浸入采样液中混匀；
    步骤四，将装置放置于水平台面，将采样液全部加入加样孔中，盖上加样孔盖；
    步骤五，反应温度50-75℃，静置10-30分钟；
    步骤六，随即将清洗液加入加样孔中，盖上加样孔盖，静置5-15分钟；
    步骤七，读取结果。
  16. 根据权利要求1所述的一种自驱动微流控检测装置的用途，其特征在于，用于肛拭子中肠炎沙门氏菌检测包括如下内容：
    采用2通道自驱动微流控检测装置，2通道自驱动微流控检测装置为加样孔通过毛细管导流渠与两个功能腔室连接，每个功能腔室对应一条层析试纸，一个结果阅读窗；
    所述毛细管导流渠的毛细管导流片的材质为玻璃材质，毛细管导流片之间的间距为0.5mm；
    采用的层析试纸为核酸免疫层析试纸；
    步骤一，将特异性检测肠炎沙门氏菌的扩增试剂系统，装有人缘GAPDH基因的质控扩增试剂系统预装于2个功能腔室中；
    所述肠炎沙门氏菌的扩增试剂系统的引物探针包括：
    正向外围引物：CGTGATGCTGAAAGTACCGA，
    反向外围引物：GGCCGCCAAAACTTTCCTGA，
    正向交叉引物：CCACCGCGTACGGACTTCACCGAAACACAAACGGGCAAG，
    反向加速引物：AGATCTTTTAGCAATTGCTTCT，
    反向探针1：荧光标记物-TGCCGCGCATACGGAACAG，
    反向探针2：荧光标记物-CCACCGCGTACGGACTTCAC；
    所述装有人缘GAPDH基因的质控扩增试剂系统的引物探针包括：
    正向外围引物：AGAACGGGAAGCTTGTCATC，
    反向外围引物：CGAACATGGGGGCATCAG，
    正向交叉引物：CAGAGGGGGCAGAGATGAATCTTCCAGGAGCGAGATCC，
    反向加速引物：ATCTTCCAGGAGCGAGATCCCAGAGGGGGCAGAGATGA，
    反向探针1：荧光标记物-CAAAATCAAGTGGGGCGA，
    反向探针2：荧光标记物-GGGAGCCAAAAGGGTC；
    步骤二，采集肛拭子；
    步骤三，将拭子头部浸入采样液中混匀，使用滤膜对采样液进行过滤，滤去残渣；
    步骤四，将装置放置于水平台面，将采样液全部加入加样孔中，盖上加样孔盖；
    步骤五，反应温度50-75℃，静置10-30分钟；
    步骤六，随即将清洗液加入加样孔中，盖上加样孔盖，静置5-15分钟；
    步骤七，读取结果。
  17. 根据权利要求1所述的一种自驱动微流控检测装置的用途，其特征在于，用于尿液样本中微生物检测包括如下内容：
    采用4通道自驱动微流控检测装置，4通道自驱动微流控检测装置为加样孔通过毛细管导流渠与四个功能腔室连接，每个功能腔室对应一条层析试纸，一个结果阅读窗。；
    采用的层析试纸为核酸免疫层析试纸；
    所述毛细管导流渠的毛细管导流片的材质为PE材质，毛细管导流片之间的间距为0.5mm；
    步骤一，将大肠杆菌检测试剂系统，肠炎沙门氏菌检测试剂系统预装于4个功能腔室中；
    所述解脲支原体检测试剂系统的引物探针包括：
    正向外围引物：GTGATTTAACTGTAGAACAAGAACA，
    反向外围引物：AGGACCACTATATTGTAGTAGTGC，
    正向交叉引物：GGCATGCGATATGAAACACCATAGATCTTTTTTGACCAGGATC，
    反向加速引物：ATTATGATTTTTAACTGGTTCTTC，
    反向探针1：荧光标记物-CACCATTTTTAATTACAGTAACT，
    反向探针2：荧光标记物-GGCATGCGATATGAAACACCA；
    所述金黄色葡萄球菌检测试剂系统的引物探针包括：
    正向外围引物：CTGAATATGCAATGAAAGTAACTGA，
    反向外围引物：TTTTTCTCTTTGCATATTATCGC，
    正向交叉引物：GACAACGCTTCTTTATCATTTGTGACAAGAGCTAGAGTCGTTAGC，
    反向加速引物：ATAATTTCTTCAAGTCGTGCCGC，
    反向探针1：荧光标记物-GTGATACCAGCATGAATCGGTTTA，
    反向探针2：荧光标记物-GACAACGCTTCTTTATCATTTGTGA；
    所述淋病奈瑟球菌检测试剂系统的引物探针包括：
    正向外围引物：GCTTTTAAATCCAATACCGTATT，
    反向外围引物：TTGAGTTCGATGGTGCTG，
    正向交叉引物：GAGGCCATTTACGCCCAATCAACAATAAAATATCCATCACCACTG，
    反向加速引物：GTGCCGTCAAGGGAAGGTTG，
    反向探针1：荧光标记物-GCCCAATCCCAAGCCGTCG，
    反向探针2：荧光标记物-GAGGCCATTTACGCCCAATC；
    所述白色念球菌检测试剂系统的引物探针包括：
    正向外围引物：CGAGTTGCCCCAAGACATG，
    反向外围引物：AATGACCGCTCTGAGTGATG，
    正向交叉引物：CAGGCCACAAACCCACCAAAGAGAATTGTCGAAAATCGCCCG，
    反向加速引物：GTGCTCTAATGGGGCAATTTCCA，
    反向探针1：荧光标记物-ATGCTGAGCCGGAGCCTTTA，
    反向探针2：荧光标记物-CAGGCCACAAACCCACCAAAGA；
    步骤二，使用干燥的拭子头部沾取尿液样本，当拭子顶部吸收到样本即可取出；
    步骤三，将拭子头部浸入采样液中混匀；
    步骤四，将装置放置于水平台面，将采样液全部加入加样孔中，盖上加样孔盖；
    步骤五，反应温度50-75℃，静置10-30分钟；
    步骤六，随即将清洗液加入加样孔中，盖上加样孔盖，静置5-15分钟；
    步骤七，读取结果。
  18. 根据权利要求1所述的一种自驱动微流控检测方法，其特征在于，用于定量检测肾脏标志物包括如下内容：
    采用4通道自驱动微流控检测装置，4通道自驱动微流控检测装置为加样孔通过毛细管导流渠与四个功能腔室连接，每个功能腔室对应一条层析试纸，一个结果阅读窗；
    所述毛细管导流渠的毛细管导流片的材质为PP材质，毛细管导流片之间的间距为0.4mm；
    步骤一，制备纳米金；
    步骤二，制备金标RBP单克隆抗体1；
    步骤三，将NC膜附在PVC底板，用往复划膜仪在NC膜的各个位置喷RBP单克隆抗体2及羊抗鼠二抗，分别为检测线(T线)和质控线(C线)，烘干；
    步骤四，在金标RBP单克隆抗体溶液中加入细胞融合剂；然后喷于玻璃纤维膜的结合垫上，烘干；
    步骤五，将玻璃纤维素膜的样品垫、玻璃纤维素膜的结合垫、NC膜和吸水垫组装成为检测试纸条，调整试纸条的灵敏度；
    步骤六，将检测试纸条组装于自驱动微流控检测装置中；
    步骤七，在4个功能腔室中依次预装四个梯度的生理盐水；
    步骤八，将尿液样本加入检测装置的加样孔中，吸出多余的样本，在微流控导流装置的控制下，进入4个功能腔室中，样本在4个功能腔室中分别被稀释100倍，10倍稀释，2倍稀释，和不稀释。
    步骤九，读取结果。
  19. 根据权利要求1所述的一种自驱动微流控检测装置的用途，其特征在于，应用于甲流病毒、乙流病毒检测包括如下内容：
    采用2通道自驱动微流控检测装置，2通道自驱动微流控检测装置为加样孔通过毛细管导流渠与两个功能腔室连接，每个功能腔室对应一条层析试纸，一个结果阅读窗；在核酸免疫试纸上设置有两条T线，分别显示甲流和乙流的检测结果；
    采用的层析试纸为核酸免疫层析试纸；
    所述毛细管导流渠的毛细管导流片的材质为玻璃材质，毛细管导流片之间的间距为0.2mm；
    步骤一，将甲流病毒检测试剂系统、将乙流病毒检测试剂系统，按照系统的成份组成，依次预装于两个功能腔室中；
    所述甲流病毒检测试剂系统的引物探针包括：
    正向外围引物：CAGAGGGCAATGATGGATCA，
    反向外围引物：ATCCCGACCAGTGAGTACC，
    正向交叉引物：CCTCAGAATGAGTGCTGACCGTAAGTCGAAACCCAGGAAACG，
    反向加速引物：CTTCCCTTTCAAAGTCATGCCCA，
    反向探针1：荧光标记物-AGGAAAATGAGGTCTTCAATCTCAG，
    反向探针2：荧光标记物-CCTCAGAATGAGTGCTGACCGT；
    所述乙流病毒检测试剂系统的引物探针包括：
    正向外围引物：ACTTACCAATGGGTGCTTAA，
    反向外围引物：CGAAAAACAGAAAGGCAACAA，
    正向交叉引物：GCATCCCATTGGAACATGTCTTCAAATTTAGTAACATTGAAGGCTCAG，
    反向加速引物：CCTCAGAAGATGGCTGGTCAGTTTTCATAACCTCTTGGTCTC，
    反向探针1：荧光标记物-GTCTTCTTTTCCCAAAAGAAACTG，
    反向探针2：荧光标记物-CAAGAGCAGTGCTCAAACAAATGA；
    步骤二，采集测试者咽拭子样本；
    步骤三，将拭子头部浸入采样液中混匀；
    步骤四，将装置放置于水平台面，将采样液全部加入加样孔中，盖上加样孔盖；
    步骤五，反应温度50-65℃，静置10-30分钟；
    步骤六，随即将清洗液加入加样孔中，盖上加样孔盖，静置5-15分钟；
    步骤七，读取结果。
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