CN111925925B - Integrated diagnostic kit and application thereof - Google Patents
Integrated diagnostic kit and application thereof Download PDFInfo
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- CN111925925B CN111925925B CN202010826506.3A CN202010826506A CN111925925B CN 111925925 B CN111925925 B CN 111925925B CN 202010826506 A CN202010826506 A CN 202010826506A CN 111925925 B CN111925925 B CN 111925925B
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- 238000009007 Diagnostic Kit Methods 0.000 title claims abstract description 16
- 238000003860 storage Methods 0.000 claims abstract description 123
- 238000004140 cleaning Methods 0.000 claims abstract description 99
- 239000007788 liquid Substances 0.000 claims abstract description 96
- 230000003321 amplification Effects 0.000 claims abstract description 51
- 238000003199 nucleic acid amplification method Methods 0.000 claims abstract description 51
- 239000003480 eluent Substances 0.000 claims abstract description 46
- 238000005336 cracking Methods 0.000 claims abstract description 45
- 239000002699 waste material Substances 0.000 claims abstract description 30
- 239000011324 bead Substances 0.000 claims abstract description 29
- 238000010828 elution Methods 0.000 claims abstract description 19
- 238000001514 detection method Methods 0.000 claims abstract description 12
- 108090000623 proteins and genes Proteins 0.000 claims abstract description 8
- 238000000197 pyrolysis Methods 0.000 claims abstract description 7
- 230000009471 action Effects 0.000 claims abstract description 5
- 239000006166 lysate Substances 0.000 claims description 29
- 238000005406 washing Methods 0.000 claims description 23
- 238000000034 method Methods 0.000 claims description 15
- 239000003153 chemical reaction reagent Substances 0.000 claims description 14
- 238000006243 chemical reaction Methods 0.000 claims description 13
- 238000003756 stirring Methods 0.000 claims description 8
- 238000012408 PCR amplification Methods 0.000 claims description 7
- 238000004108 freeze drying Methods 0.000 claims description 6
- 230000009089 cytolysis Effects 0.000 claims description 4
- 239000012530 fluid Substances 0.000 claims 1
- 238000003745 diagnosis Methods 0.000 abstract description 4
- 241000700605 Viruses Species 0.000 abstract description 3
- 208000024891 symptom Diseases 0.000 abstract description 3
- 239000003814 drug Substances 0.000 abstract description 2
- 244000052616 bacterial pathogen Species 0.000 abstract 1
- 239000000047 product Substances 0.000 description 9
- 230000008569 process Effects 0.000 description 6
- 239000012634 fragment Substances 0.000 description 4
- 238000013459 approach Methods 0.000 description 2
- 230000009286 beneficial effect Effects 0.000 description 2
- 201000010099 disease Diseases 0.000 description 2
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 2
- 238000007403 mPCR Methods 0.000 description 2
- 244000052769 pathogen Species 0.000 description 2
- 238000012360 testing method Methods 0.000 description 2
- 208000035473 Communicable disease Diseases 0.000 description 1
- 230000004075 alteration Effects 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 230000008901 benefit Effects 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 238000003776 cleavage reaction Methods 0.000 description 1
- 238000004891 communication Methods 0.000 description 1
- 230000002860 competitive effect Effects 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 239000012502 diagnostic product Substances 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
- 231100000676 disease causative agent Toxicity 0.000 description 1
- 238000005538 encapsulation Methods 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 238000000605 extraction Methods 0.000 description 1
- 208000015181 infectious disease Diseases 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 238000012123 point-of-care testing Methods 0.000 description 1
- 238000001179 sorption measurement Methods 0.000 description 1
- 238000006467 substitution reaction Methods 0.000 description 1
- 229920001169 thermoplastic Polymers 0.000 description 1
- 239000004416 thermosoftening plastic Substances 0.000 description 1
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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
- C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
- C12Q1/6806—Preparing nucleic acids for analysis, e.g. for polymerase chain reaction [PCR] assay
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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
- C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
- C12Q1/6844—Nucleic acid amplification reactions
- C12Q1/686—Polymerase chain reaction [PCR]
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- Proteomics, Peptides & Aminoacids (AREA)
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Abstract
The invention discloses an integrated diagnostic kit and application thereof, and relates to the technical field of life medicine detection and diagnosis. The body of the kit is disc-shaped and comprises a sample loading groove, a cleaning solution storage groove, a cracking solution storage groove, an eluent storage groove, an oil storage groove, a cracking cleaning groove, an elution groove, an amplification groove and a waste liquid storage groove; the liquid storage area comprises a pyrolysis liquid tank, a cleaning liquid tank, an eluent tank and an oil tank; the sample loading groove is positioned in the middle of the disc, the liquid storage area is distributed concentrically with the sample loading groove, and the amplification groove is positioned at the edge of the kit. According to the invention, by designing the disc structure, the matching motor drives the kit to rotate to generate centrifugal force, and under the action of the magnetic force and the PCR magnetic beads, the sample genes are subjected to cracking cleaning, eluting and amplifying, and finally the detection is carried out to obtain a result. The invention shortens the detection time to 20 to 30 minutes by adopting a disc structure, and can detect various germs and viruses causing symptoms simultaneously.
Description
Technical Field
The invention relates to the technical field of life medicine detection and diagnosis, in particular to an integrated diagnosis kit and application thereof.
Background
In the field of infectious diseases, in order to control the spread of infection, it is necessary to rapidly derive the causative agent, or whether personnel are infected, and very rapid and accurate molecular diagnostic products are required.
The FilmArray product of BioFire is a classic task that has been successfully commercialized at present, and uses nested multiplex PCR analysis techniques. The whole FilmArray reagent can be simply divided into two parts, an upper reservoir set and a lower reaction layer. The basic structure of such a tube set and the principle and procedure of drawing liquid using vacuum are disclosed in patent US8409508, where a single tube is attached to a flexible bag by means of a thermoplastic seal, wherein the bottom of the tube is in communication with a reaction tank within the flexible bag. The liquid storage tube wall has two openings, the upper opening is the gas vent, is used for the inside air formation vacuum cavity of extraction when liquid storage tube encapsulation, and the lower opening is reagent entry, generally is sealed state, and reagent can be inhaled by the vacuum after puncturing.
As with other POCT products, the FilmArray is also detected by adopting a mode of a testing instrument and a disposable testing chip, and compared with other disease detection chips on the market, the FilmArray has very obvious competitive advantage, can detect various pathogens and viruses causing symptoms at the same time, and finally locates the cause of the disease, and is mainly beneficial to the multiple PCR technology.
However, the manufacturing process of the kit is very complex, so that the product price of the FilmArray is high, and therefore, an integrated diagnosis kit is necessary to be specifically designed, the problem of process complexity is solved on the premise of meeting various detection requirements and not changing the performance and convenience of the product, and the cost of the product is reduced.
Disclosure of Invention
Aiming at the defects of the prior art, the invention provides an integrated diagnostic kit, which solves the problem of process complexity and reduces the cost of products on the premise of meeting various detection requirements and not changing the performance and convenience of the products.
In order to achieve the above purpose, the invention provides an integrated diagnostic kit, which comprises a disc-shaped kit body, and a sample loading groove, a cleaning solution storage groove, a lysate storage groove, an eluent storage groove, an oil storage groove, a lysis washing groove, an elution groove, an amplification groove and a waste liquid storage groove which are distributed on the body;
wherein:
the sample loading groove is positioned in the middle of the kit, and the cleaning liquid storage groove, the cracking liquid storage groove, the eluent storage groove and the oil storage groove are concentrically arranged with the sample loading groove; the amplification tank is positioned close to the edge of the kit, and the lysis washing tank and the elution tank are close to the liquid storage area;
the sample loading groove is communicated with the cracking cleaning groove through a sample loading flow channel; the cracking cleaning tank is communicated with the waste liquid storage tank through a cleaning waste liquid runner; the cracking cleaning tank is communicated with the elution tank through a magnetic bead moving channel; the elution groove is communicated with the amplification groove through a capillary flow passage;
at least one of the cleaning liquid storage tank, the cracking liquid storage tank, the eluent storage tank and the oil storage tank;
the cleaning solution storage tank is provided with a cleaning solution overflow port, the cleaning solution overflow port is communicated with a cleaning solution flow channel, and the cleaning solution flow channel is communicated with the cracking cleaning tank;
the cracking liquid storage tank is provided with a cracking liquid overflow port, and the cracking liquid overflow port is communicated with the cracking cleaning tank;
the eluent storage tank is provided with an eluent overflow port, and the eluent overflow port is communicated with the eluent storage tank;
the oil storage tank is provided with an amplification flow channel which is convenient for oil to flow out, and the amplification flow channel is communicated with the amplification tank.
Further, the sample loading groove is round,
further, the sample adding groove is positioned at the center of the kit body;
further, the cleaning solution storage tank, the cracking solution storage tank, the eluent storage tank and the oil storage tank are arranged around the sample loading tank and are all fan-shaped;
further, the cleaning solution storage tanks are provided with more than two overflow ports, the cleaning solution storage tanks are respectively communicated with a cleaning solution runner, and the cleaning solution runner is annular and is positioned on the outer wall of the cleaning solution storage tanks;
further, the amplification tank comprises more than two amplification tanks which are communicated in sequence; the amplification groove and the sample loading groove are concentrically arranged;
further, the waste liquid storage groove and the sample loading groove are concentrically arranged and are close to the edge of the kit;
further, the cleaning waste liquid flow channel is U-shaped or S-shaped;
further, the capillary flow passage is U-shaped or S-shaped.
It is still another object of the present invention to provide a method for detecting a target gene using an integrated diagnostic kit, comprising the steps of:
1. placing the kit on a matched workbench with a motor, adding a PCR sample into a sample adding groove, connecting the kit with the matched motor, adding a lysate into a lysate storing groove, starting the motor, synchronously rotating the kit, allowing the sample and the lysate to enter a lysate washing groove under the action of centrifugal force, adding PCR magnetic beads, and performing a cleavage reaction on the sample; after the reaction, adding magnetic force, and centrifugally flowing the reacted pyrolysis liquid into a waste liquid storage tank by the centrifugal force of a motor;
2. removing the magnetic force, adding cleaning liquid into the cleaning liquid storage tank, starting the motor, stopping the motor after the cleaning liquid flows into the cracking cleaning tank, and cleaning under the vibration stirring; after the cleaning is finished, magnetic force is applied, and the aggregated magnetic beads enter an elution groove along a magnetic bead channel; starting a motor, and centrifugally flowing the cleaning waste liquid into a waste liquid storage tank;
3. adding eluent into the eluent storage tank, rotating along with a motor to enter the eluent tank, removing magnetic force, stopping the motor, performing vibration stirring elution operation, adding magnetic force to remove magnetic beads after the completion of the vibration stirring elution operation, starting the motor, and enabling the eluent to enter the amplification tank through a capillary flow passage;
4. adding oil into the oil storage tank, entering the amplification tank along with the rotation of the motor, controlling the temperature-changing circulation of the cavity of the motor to perform amplification reaction, and detecting fluorescent signals of the amplification tank after the reaction is finished to obtain a detection result.
Preferentially, the step 2 can be repeated for two or more times as required;
further, the freeze-drying reagent required by PCR amplification is stored in the amplification tank in the step 4 in advance, and the freeze-drying reagent comprises primers required by the amplification reaction and other components required by the PCR amplification;
further, the step 4 may be performed a plurality of times of amplification as needed.
The working principle of the integrated diagnostic kit provided by the invention is as follows: under the action of the rotating centrifugal force of the matched motor, corresponding reagents in the kit can flow into the communicated areas along with the overflow port and respectively enter the cracking washing tank, the eluting tank and the amplifying tank in sequence, the cracking washing and eluting are completed by means of magnetic force adsorption or release of PCR magnetic beads, and finally the reagents enter the amplifying tank for amplification and reaction, and the amplified fluorescent signals are detected to obtain detection results.
Compared with the prior art, the invention has the following beneficial effects:
1. the scheme of the invention adopts a disc structure, so that the problem of process complexity can be solved, the cost of the product is greatly reduced on the premise of not changing the performance and convenience of the product, and the detection time is shortened to 20 to 30 minutes;
2. the kit is used for multiplex PCR amplification, and can be used for simultaneously detecting various pathogens and viruses causing symptoms.
Drawings
FIG. 1 is a schematic diagram of one embodiment of an integrated diagnostic kit of the present invention.
FIG. 2 is an enlarged schematic view of a portion of one embodiment of an integrated diagnostic kit of the present invention.
In the figure: 1 sample loading tank, 2 washing liquid storage tank, 3 lysate storage tank, 4 eluent storage tank, 5 oil storage tank, 6 lysate washing tank, 7 elution tank, 8 amplification tank, 9 waste liquid storage tank, 10 washing waste liquid runner, 11 capillary runner, 12 lysate overflow port, 13 loading runner, 14 washing liquid overflow port, 15 washing liquid runner, 16 magnetic bead moving channel, 18 amplification runner, 19 reagent box body.
Detailed Description
The following description of the embodiments of the present invention will be made clearly and completely with reference to the accompanying drawings, in which it is apparent that the embodiments described are only some embodiments of the present invention, but not all embodiments. All other embodiments, which can be made by those skilled in the art based on the embodiments of the invention without making any inventive effort, are intended to be within the scope of the invention.
Examples
Referring to fig. 1 and 2 (fig. 2 is an enlarged view of a portion a in fig. 1), the embodiment provides an integrated diagnostic kit, wherein a kit body (19) is disc-shaped, and the integrated diagnostic kit comprises: a sample loading tank (1), a liquid storage area consisting of a cleaning solution storage tank (2), a cracking solution storage tank (3), an eluent storage tank (4) and an oil storage tank (5), a cracking cleaning tank (6), an elution tank (7), an amplification tank (8) and a waste liquid storage tank (9);
the sample adding groove (1) is circular and is positioned in the middle part, preferably the center position, of the kit; the liquid storage area is concentrically arranged around the sample application well (1); the positions of the amplification tank (8) and the waste liquid storage tank (9) are close to the edge of the kit; at least one amplification groove (8) (5 in the embodiment and the attached drawings) is communicated in sequence, is concentric with the sample adding groove (1) and is arranged in a fan shape, and the number of the amplification grooves can be adjusted according to the requirement; the waste liquid storage groove (9) is concentrically arranged with the sample adding groove (1); the cracking cleaning tank (6) and the eluting tank (7) are close to the liquid storage area;
a sample adding flow channel (13) is arranged between the sample adding groove (1) and the cracking cleaning groove (6), and after the sample is added into the sample adding groove (1), the sample can enter the cracking cleaning groove (6) through the sample adding flow channel (13); a cleaning waste liquid flow channel (10) is arranged between the cracking cleaning tank (6) and the waste liquid storage tank (9); a magnetic bead moving channel (16) is arranged between the cracking washing tank (6) and the eluting tank (7); a capillary flow passage (11) is arranged between the elution groove (7) and the amplification groove (8); the cleaning waste liquid flow channel (10) and the capillary flow channel (11) are both U-shaped (the structures of the embodiment and the drawing are both U-shaped; one of the two can be designed into an S-shape or an S-shape according to actual needs);
the liquid storage area comprises at least one of a cleaning liquid storage tank (2), a cracking liquid storage tank (3), an eluent storage tank (4) and an oil storage tank (5) (in the embodiment and the drawing, the number of the cleaning liquid storage tanks is 4, the number of the cracking liquid storage tanks is 1, the number of the eluent storage tanks is 1, the number of the oil storage tanks can be actually adjusted according to the needs of each program), and the cleaning liquid storage area is fan-shaped (the embodiment and the drawing are fan-shaped structures, and can be designed into other various shapes);
a cleaning liquid overflow port (14) is arranged on the cleaning liquid storage tank (2), and the cleaning liquid overflow port (14) is communicated with a cleaning liquid flow channel (15); the cleaning solution runner (15) is positioned on the outer wall of the cleaning solution storage tank and is communicated with the cracking cleaning tank (6);
a pyrolysis liquid overflow port (12) is arranged on the pyrolysis liquid storage tank (3), and the pyrolysis liquid overflow port (12) is communicated with the pyrolysis cleaning tank (6);
an eluent overflow port (17) is arranged on the eluent storage tank (4), and the eluent overflow port (17) is communicated with the eluent tank (7);
the oil storage tank (5) is provided with an amplification flow channel (18) which is convenient for oil to flow out, and the amplification flow channel (18) is communicated with the amplification tank (8).
For the cleaning liquid overflow port (14), the lysate overflow port (12), the eluent overflow port (17) and the amplification flow channel (18), the overflow port is arranged on one arc side of the corresponding storage tank in the embodiment and the drawing, so that the liquid can flow out conveniently.
The sample loading groove (1), the cleaning solution storage groove (2), the lysate storage groove (3), the eluent storage groove (4) and the oil storage groove (5) are grooves, and gaps are formed between adjacent grooves or not; the same cleaning liquid storage tank (2) contains or does not contain gaps;
in this embodiment, gaps are included or not included between the amplification cells (8).
The working principle of the integrated diagnostic kit is described by combining the following using method:
a) The kit is placed on a matched workbench with a motor, a PCR sample is added into a sample adding groove (1), then the kit is placed into a detection instrument, and the kit is connected with the motor to form a whole. The lysate is added into the lysate storage tank (3) (in this embodiment, the lysate can be pre-packaged in the lysate bag and placed in the lysate storage tank, when in use, the liquid bag is opened by puncturing or extruding to release the liquid), the motor is started to drive the reagent box to rotate, under the action of centrifugal force, the sample and the lysate respectively enter the lysate washing tank (6) from the sample adding flow channel (13) and the lysate overflow port (12), and the magnetic beads in the PCR are pre-placed in the lysate washing tank or the lysate and also enter the same. At the moment, a sample is arranged in the cracking washing tank (6), the cracking liquid and the magnetic beads are dispersed in an oscillating and stirring mode, and the gene fragments are adsorbed by the magnetic beads after the sample is fully cracked; a permanent magnet or an electromagnet is arranged below the cracking washing tank (6), after the gene fragments are fully adsorbed, the permanent magnet is lifted or the electromagnet is electrified to generate magnetic force on magnetic beads in the cracking washing tank (6), the magnetic beads are clustered and approach to the bottom of the cracking washing tank (6), and the motor rotates to convey the cracking liquid in the cracking washing tank (6) into a waste liquid storage tank (9) through a cleaning waste liquid runner (10).
b) The cleaning solution storage tank (2) is added with cleaning solution (in the embodiment, the cleaning solution can be packaged in the cleaning solution storage tank in the cleaning solution bag in advance, the liquid bag is opened to release the liquid in a puncturing or squeezing way when in use), the centrifugal force of the motor conveys the cleaning solution from the cleaning solution overflow port (14) to the cracking cleaning tank (6), the permanent magnet descends or the electromagnet is powered off, and the magnetic beads are dispersed in a mode of driving the reagent box to vibrate and stir by the motor so as to be fully contacted with the cleaning solution for cleaning; the permanent magnet or the electromagnet is electrified again, the magnetic beads are gathered and approach to the bottom, and the motor centrifugal force conveys the cleaning waste liquid into the waste liquid storage tank (9) through the cleaning waste liquid flow channel (10) to complete a cleaning flow. The cleaning process can be repeated for a plurality of times according to the requirement, so as to meet the cleaning requirement.
c) After the washing step is finished, magnetic beads are adsorbed and moved by magnetic force, the magnetic beads enter an elution tank (7) through a magnetic bead moving channel (16), eluent is added into an eluent storage tank (4) (in the embodiment, the eluent can be packaged in an eluent bag in advance and is placed in the eluent storage tank, when the eluent storage tank is used, liquid is released by opening a liquid bag in a puncturing or squeezing mode), the centrifugal force of a motor drives the eluent to enter the elution tank (7) from an eluent overflow port (17), the magnetic beads are released by a magnet, and the magnetic beads are subjected to vibration stirring, dispersed and fully mixed with the eluent for elution; and (3) starting magnetic force again to gather the magnetic beads, moving the magnet to return the magnetic beads to the lysis washing tank (6) through the magnetic bead moving channel (16), starting the motor at the moment, and enabling the eluent to enter the amplification tank (8) through the capillary flow channel (11) with the gene fragments.
d) The oil is added into the oil storage tank (5), the motor rotates, the oil enters the amplification tank (8) from the amplification flow channel (18) (in the embodiment, the oil can be packaged in the oil storage tank in advance, and released by opening the liquid package in a puncturing or extruding mode when in use), at this time, the motor continuously rotates, and the amplification tank (8) is controlled by temperature rise and fall, so that the PCR amplification system circulates by temperature rise and fall. And detecting fluorescent signals of the amplification groove after the temperature change cycle is finished to determine whether target gene fragments exist in the sample. Wherein, according to whether multiple PCR is needed, the number of the amplifying tanks (8) is set, freeze-drying reagent components needed by PCR amplification are stored in the amplifying tanks (8) in advance, and the freeze-drying reagent components comprise primers needed by the amplification reaction and other components needed by the PCR amplification, and the primers in different amplifying tanks (8) are different.
It should be noted that, in this document, the term "comprises," "comprising," or any other variation thereof, are intended to cover a non-exclusive inclusion, such that a process, method, article, or apparatus that comprises a list of elements does not include only those elements but may include other elements not expressly listed or inherent to such process, method, article, or apparatus.
Although embodiments of the present invention have been shown and described, it will be understood by those skilled in the art that various changes, modifications, substitutions and alterations can be made therein without departing from the principles and spirit of the invention, the scope of which is defined in the appended claims and their equivalents.
Claims (6)
1. An integrated diagnostic kit is characterized by comprising a disc-shaped kit body (19), and a sample loading groove (1), a cleaning liquid storage groove (2), a lysate storage groove (3), an eluent storage groove (4), an oil storage groove (5), a lysate washing groove (6), an elution groove (7), an amplification groove (8) and a waste liquid storage groove (9) which are distributed on the kit body (19);
the sample adding groove (1) is positioned in the middle of the kit; the cleaning solution storage tank (2), the lysate storage tank (3), the eluent storage tank (4) and the oil storage tank (5) are concentrically arranged with the sample loading tank (1); the amplification tank (8) is positioned close to the edge of the kit, and the lysis rinsing tank (6) and the elution tank (7) are close to the liquid storage area;
the sample loading groove (1) is communicated with the cracking cleaning groove (6) through a loading flow channel (13); the cracking cleaning tank (6) is communicated with the waste liquid storage tank (9) through a cleaning waste liquid flow channel (10); the cracking cleaning tank (6) is communicated with the eluting tank (7) through a magnetic bead moving channel (16); the elution groove (7) is communicated with the amplification groove (8) through a capillary flow passage (11);
at least one of the cleaning solution storage tank (2), the lysate storage tank (3), the eluent storage tank (4) and the oil storage tank (5);
the cleaning solution storage tank (2) is provided with a cleaning solution overflow port (14), and the cleaning solution overflow port (14) is communicated with a cleaning solution flow channel (15); the cleaning fluid flow channel (15) is communicated with the cracking cleaning tank (6);
the cracking liquid storage tank (3) is provided with a cracking liquid overflow port (12), and the cracking liquid overflow port (12) is communicated with the cracking cleaning tank (6);
the eluent storage tank (4) is provided with an eluent overflow port (17), and the eluent overflow port (17) is communicated with the eluent tank (7);
the oil storage tank (5) is provided with an amplification flow channel (18) which is convenient for oil to flow out, and the amplification flow channel (18) is communicated with the amplification tank (8);
the sample adding groove (1) is round and is positioned in the center of the kit body (19);
the cleaning solution storage tank (2), the cracking solution storage tank (3), the eluent storage tank (4) and the oil storage tank (5) are arranged around the sample loading tank (1) and are all fan-shaped;
the amplification groove (8) comprises more than two and is sequentially communicated with the sample adding groove (1) in a concentric manner;
the waste liquid storage groove (9) is concentrically arranged with the sample loading groove (1) and is close to the edge of the kit.
2. The integrated diagnostic kit according to claim 1, wherein the cleaning liquid storage tank (2) comprises more than two cleaning liquid overflow ports (14) are respectively arranged on the cleaning liquid storage tank (2), the cleaning liquid overflow ports (14) are respectively communicated with a cleaning liquid flow channel (15), and the cleaning liquid flow channel is annular and is positioned on the outer wall of the cleaning liquid storage tank (2).
3. The integrated diagnostic kit according to claim 1, wherein the washing waste liquid flow channel (10) is U-shaped or S-shaped; the capillary flow passage (11) is U-shaped or S-shaped.
4. A method for detecting a target gene using the integrated diagnostic kit of claim 1, for non-diagnostic purposes, comprising the steps of:
s1, placing the kit on a matched workbench with a motor, adding a PCR sample into a sample adding groove (1), adding a lysate into a lysate storing groove (3), starting the motor, synchronously rotating the kit, enabling the sample and the lysate to enter a lysate washing groove (6) under the action of centrifugal force, adding PCR magnetic beads, and carrying out a lysate reaction on the sample; after the reaction is finished, magnetic force is added, and the reacted pyrolysis liquid centrifugally flows into a waste liquid storage tank (9) by the centrifugal force of a motor;
s2, removing the magnetic force, adding cleaning liquid into the cleaning liquid storage tank (2), starting the motor, stopping the motor after the cleaning liquid flows into the cracking cleaning tank (6), and cleaning under the vibration and stirring; after the cleaning is finished, magnetic force is applied, and the aggregated magnetic beads enter an elution groove (7) along a magnetic bead moving channel (16); starting the motor, and centrifugally flowing the cleaning waste liquid into a waste liquid storage tank (9);
s3, adding eluent into the eluent storage tank (4), rotating along with a motor to enter the eluent storage tank, removing magnetic force, stopping the motor, performing vibration stirring elution operation, removing magnetic beads by adding the magnetic force after completion, starting the motor, and allowing the eluent to enter the amplification tank (8) through the capillary flow channel (11);
and S4, adding oil into the oil storage tank (5), entering the amplification tank (8) along with the rotation of the motor, controlling the temperature-changing circulation of the cavity to perform amplification reaction, and detecting fluorescent signals of the amplification tank after the reaction is finished to obtain a detection result.
5. The method for detecting a target gene according to claim 4, wherein the step S2 is repeated two or more times.
6. The method according to claim 4, wherein the freeze-drying reagent required for PCR amplification is stored in the amplification tank (8) in step S4, and the freeze-drying reagent contains the primers required for the amplification reaction and other components required for PCR amplification.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202010826506.3A CN111925925B (en) | 2020-08-17 | 2020-08-17 | Integrated diagnostic kit and application thereof |
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| Application Number | Priority Date | Filing Date | Title |
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| CN202010826506.3A CN111925925B (en) | 2020-08-17 | 2020-08-17 | Integrated diagnostic kit and application thereof |
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| Publication Number | Publication Date |
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| CN111925925A CN111925925A (en) | 2020-11-13 |
| CN111925925B true CN111925925B (en) | 2024-02-27 |
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Citations (11)
| Publication number | Priority date | Publication date | Assignee | Title |
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