CN210394313U - Fan-shaped nucleic acid multi-joint detection device - Google Patents
Fan-shaped nucleic acid multi-joint detection device Download PDFInfo
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- CN210394313U CN210394313U CN201920387799.2U CN201920387799U CN210394313U CN 210394313 U CN210394313 U CN 210394313U CN 201920387799 U CN201920387799 U CN 201920387799U CN 210394313 U CN210394313 U CN 210394313U
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Abstract
The utility model discloses a fan-shaped nucleic acid allies oneself with examines device more, include: the device comprises a disc type centrifugal support, a fan-shaped chip and a closed film, wherein the disc type centrifugal support is connected with the fan-shaped chip, and the fan-shaped chip is connected with the closed film; the device can pre-embed the primer probe of different projects in each reaction hole respectively, can realize the detection of disposable multiple projects, consumes to hang down, simple operation, the leakproofness is good, be difficult for producing the aerosol pollution.
Description
Technical Field
The utility model relates to the field of biotechnology, concretely relates to fan-shaped nucleic acid allies oneself with examines device more.
Background
The PCR technology emerged in 1985, and with the development of more than thirty years, the PCR technology has been rapidly developed and widely applied, and particularly in clinical diagnosis of diseases, the PCR technology increasingly replaces the traditional microorganism diagnosis method with the advantages of rapidness, sensitivity and specificity, and enters conventional detection. Many nucleic acid Amplification technologies derived from the PCR technology include real-time fluorescence quantitative PCR, loop-Mediated isothermal Amplification (LAMP), and normal temperature nucleic acid Amplification (EMA).
Among them, the real-time fluorescent quantitative PCR technology is the most widely used technology, and many companies in China develop research works for detecting products aiming at the technology. The products mainly use 200 mul PCR tubes, a single tube mainly detects a single index, the reaction volume is 20-50 mul, and the price of a matched fluorescence quantitative instrument is expensive, so that wastes such as samples, reagents, consumables, time, personnel cost, instrument maintenance and the like can be caused when multiple indexes are detected, the requirement of high-throughput detection cannot be met, and the products are not easy to popularize. Meanwhile, people have higher health consciousness and scientific literacy, and are more cautious to pathogen screening and scientific medication, so that the requirement of people on pathogen high-throughput screening is met better by obtaining a multi-index detection result through single sampling.
An Enzyme Mediated Amplification (EMA) technology developed by the company is a multi-enzyme Mediated nucleic acid Amplification technology, and can detect pathogenic nucleic acid within 30 minutes at a constant temperature of 37-45 ℃. Therefore, a fan-shaped nucleic acid multi-detection device is urgently needed, each reaction hole of the device can be pre-embedded with primer probes of different items, and a multi-index detection result can be obtained within 30 minutes after a sample is sampled at one time. The single-hole reaction volume of the device is 10 mu l, the consumption of samples, consumables and reagents is reduced, the detection time and cost are saved, the probability of missed detection of sample pathogens is reduced, and the detection flux is improved.
SUMMERY OF THE UTILITY MODEL
In order to solve the technical problem, the utility model provides a fan-shaped nucleic acid multi-link inspection device, the device each reaction hole can be pre-buried different project's primer probe respectively, can realize the detection of disposable multiple project, consumes low, the simple operation, the leakproofness is good, be difficult for producing the aerosol pollution.
In order to achieve the above purpose, the technical scheme of the utility model is as follows:
a fan-shaped nucleic acid multi-detection device comprises: the device comprises a disc type centrifugal support, a fan-shaped chip and a closed film, wherein the disc type centrifugal support is connected with the fan-shaped chip, and the fan-shaped chip is connected with the closed film.
The utility model provides a pair of fan-shaped nucleic acid multi-link inspection device, the device each reaction hole can be pre-buried different project's primer probe respectively, can realize the detection of disposable multiple nucleic acid, consumes low, simple operation, leakproofness good, be difficult for producing the aerosol pollution.
On the basis of the technical scheme, the following improvements can be made:
as a preferred scheme, a central circular truncated cone is arranged on the disc type centrifugal support and connected with the disc type centrifugal support.
As an optimized scheme, a small-sized circular truncated cone is arranged on the disc type centrifugal support and connected with the disc type centrifugal support, and the small-sized circular truncated cone is uniformly arranged along the circumference of the central circular truncated cone and is clamped with the clamping groove on one side of the fan-shaped chip.
Preferably, the disc-type centrifugal support is provided with an elastic sheet, and the other side of the fan-shaped chip is fixedly connected with the disc-type centrifugal support through the elastic sheet.
Preferably, the sector chip includes: the device comprises a liquid adding pool, a liquid separating tank, a gas displacement hole, a reaction hole and a waste liquid tank, wherein the liquid adding pool, the liquid separating tank, the gas displacement hole, the reaction hole and the waste liquid tank are connected with one another through flow channels.
Preferably, the sample is added into the liquid adding pool, the sample flows into the liquid separating groove under the action of the centrifugal force of the disc type centrifugal support, the liquid separating groove collects the sample, the gas is replaced through the gas replacing hole and the reaction hole, so that the reaction hole is filled with the sample, and the redundant sample can be discharged into the waste liquid groove.
Preferably, the sealing film is provided with a sample adding hole, the sample adding hole corresponds to and is matched with the liquid adding pool, and a sample is added into the liquid adding pool from the sample adding hole.
According to the preferable scheme, the sampling hole is provided with a sampling hole pad pasting, and the sampling hole pad pasting corresponds to and is mutually matched and connected with the sampling hole.
As a preferable scheme, six groups of medium-sized cylinders are arranged on the disc type centrifugal support.
As a preferred scheme, six groups of medium-sized cylinders are uniformly distributed on the edge of the surface of the disc-type centrifugal support, and the fan-shaped chips are placed between the two groups of medium-sized cylinders and fixedly connected with the two groups of medium-sized cylinders.
Drawings
FIG. 1 is a block diagram of a multiple nucleic acid multiplex assay device
FIG. 2 is a fan-shaped chip structure diagram of a multi-step nucleic acid detection device
FIG. 3 is a structural diagram of a sealing film in a fan-shaped nucleic acid multi-detection device
FIG. 4 is a diagram of the structure of an exhaust hole in a multiple nucleic acid detection device
FIG. 5 shows the manufacturing and testing results of the respiratory tract 5 joint inspection chip in example 3
Wherein: 1. a disc-type centrifugal support; 2. a fan-shaped chip; 3. a central right circular truncated cone; 4. a small right circular truncated cone; 5. a medium-sized cylinder; 6. a spring plate; 7. a card slot; 8. a liquid adding pool; 9. a liquid separating tank; 10. a gas displacement bore; 11. a reaction well; 12. a waste liquid tank; 13. a flow channel; 14. sealing and sticking a film; 15. a cup-shaped liquid separating tank; 16. a sample application hole; 17. pasting a film on the sampling hole; 18. and (4) exhausting holes.
Detailed Description
The technical solutions in the embodiments of the present invention will be described clearly and completely below, and it should be understood that the described embodiments are only some embodiments of the present invention, but not all embodiments. Based on the embodiments in the present invention, all other embodiments obtained by a person skilled in the art without creative efforts belong to the protection scope of the present invention.
Preferred embodiments of the present invention will be described in detail below with reference to the accompanying drawings.
In order to achieve the object of the present invention, as shown in fig. 1 to 5, a multiple nucleic acid sector inspection apparatus in the present embodiment includes: the device comprises a disc type centrifugal support 1, a fan-shaped chip 2 and a closed film 14, wherein the disc type centrifugal support 1 is connected with the fan-shaped chip 2, and the fan-shaped chip 2 is connected with the closed film 14.
The utility model provides a pair of fan-shaped nucleic acid multi-link inspection device, the device each reaction hole can be pre-buried different project's primer probe respectively, can realize the detection of disposable multiple project, consumes low, simple operation, leakproofness good, be difficult for producing the aerosol pollution.
In some embodiments, the disc-type centrifugal support 1 is provided with a central circular truncated cone 3, and the central circular truncated cone 3 is connected with the disc-type centrifugal support 1.
By adopting the embodiment, the centrifugal motion device is simple in structure, convenient to operate and convenient for centrifugal motion of the central right circular truncated cone 3.
In some embodiments, a small-sized circular truncated cone 4 is arranged on the disc-type centrifugal support 1, the small-sized circular truncated cone 4 is connected with the disc-type centrifugal support 1, and the small-sized circular truncated cone 4 is uniformly arranged along the circumference of the central circular truncated cone 3 and is clamped with a clamping groove 7 on one side of the fan-shaped chip 2.
By adopting the embodiment, the small-sized right circular truncated cone 4 is close to the central right circular truncated cone 3 and is uniformly distributed, the size of the cone angle is equal to that of the cone angle of the central right circular truncated cone 3, the bottom surface of the small-sized right circular truncated cone 4 is upward and has downward friction force, and therefore the small-sized right circular truncated cone has a buckling effect and buckles the clamping groove 7 on the small arc edge of the fan-shaped chip 2.
In some embodiments, the disc-type centrifugal support 1 is provided with a spring sheet 6, and the other side of the sector chip 2 is fixedly connected with the disc-type centrifugal support 1 through the spring sheet 6.
By adopting the embodiment, the elastic sheets 6 are uniformly distributed on the edge of the disc type centrifugal support 1 and are arranged at the outer edge of the disc type centrifugal support 1 in a protruding mode by 5 millimeters, the middle position of each 2 elastic sheets 6 corresponds to the position of the small right circular table 4, the angle a of each elastic sheet 6 is less than 80 degrees, the elastic sheets are used for fixing 1 fan-shaped chip 2, and the buffering effect is achieved.
In some embodiments, the fan chip 2 includes: the device comprises a liquid adding pool 8, a liquid separating tank 9, a gas replacement hole 10, a reaction hole 11 and a waste liquid tank 12, wherein the liquid adding pool 8, the liquid separating tank 9, the gas replacement hole 10, the reaction hole 11 and the waste liquid tank 12 are connected with one another through a flow channel 13.
By adopting the embodiment, the structure is simple and the operation is convenient.
In some embodiments, a sample is added to the cuvette 8, the sample flows into the separation tank 9 by the centrifugal force of the disk-type centrifugal support 1, the separation tank 9 collects the sample, the gas is displaced through the gas displacement holes 10 and the reaction holes 11, the reaction holes 11 are filled with the sample, and the excess sample is discharged to the waste liquid tank 12.
By adopting the embodiment, the volume of the liquid adding pool 8 is more than 60 mu l, the end, far away from the center of the circle, of the liquid adding pool 8 is connected with the liquid separating groove 9 through the flow channel 13, a sample larger than 60 mu l is firstly added into the liquid adding pool 8, the sample flows into the 5 cup-shaped liquid separating grooves 15 of the liquid separating groove 9 under the action of the centrifugal force of the disc type centrifugal support 1, and after the sample is collected by the liquid separating groove 9, the gas is displaced through the gas displacement holes 10 and the reaction holes 11, so that the 5 reaction holes 11 are filled with the sample, and redundant samples can be discharged into the waste liquid groove 12.
In some embodiments, the sealing film is provided with a sample adding hole, the sample adding hole corresponds to and is matched with the liquid adding pool, and a sample is added into the liquid adding pool from the sample adding hole.
By adopting the embodiment, the sealing adhesive film 14 covers and adheres to the upper surface of the fan-shaped chip 2, the sealing adhesive film 14 is provided with the sample adding hole 16, the sample adding hole 16 corresponds to the liquid adding pool 8, and a sample is added into the liquid adding pool 8 from the sample adding hole 16, so that the operation is convenient, and the working efficiency is improved.
In some embodiments, the sample hole 16 is provided with a sample hole pad 17, and the sample hole pad 17 corresponds to and is connected with the sample hole 16 in a matching manner.
By adopting the embodiment, the sample adding hole adhesive film 17 and the sample adding hole 16 correspond to each other and are mutually matched and connected, so that the sealing performance is improved, and aerosol pollution is not easy to generate.
In some embodiments, six sets of medium cylinders 5 are provided on the disc centrifuge rack 1.
By adopting the embodiment, the medium-sized cylinders 5 are uniformly distributed on the upper surface of the disc type centrifugal support 1 and close to the edge of the disc type centrifugal support 1, the position of each medium-sized cylinder 5 corresponds to the middle position of each 2 small-sized right circular truncated cones 4, and 1 fan-shaped chip 2 is fixed on each 2 medium-sized cylinders 5.
In some embodiments, six groups of medium cylinders 5 are uniformly distributed on the edge of the surface of the disc-type centrifugal support 1, and the fan-shaped chips 2 are placed between the two groups of medium cylinders 5 and fixedly connected with the two groups of medium cylinders 5.
By adopting the embodiment, the medium-sized cylinders 5 are uniformly distributed on the upper surface of the bracket close to the edge of the bracket, the position of each medium-sized cylinder 5 corresponds to the middle position of each 2 small right circular truncated cones 4, and each 2 medium-sized cylinders 5 fix 1 fan-shaped chip 2.
The sector nucleic acid multi-joint detection device consists of 1 disc type centrifugal support 1, 6 sector chips 2 and 6 closed adhesive films 14.
The upper surface of the disc type centrifugal support 1 is provided with 6 sectors A-F corresponding to 1 central right circular truncated cone 3, 6 concentric small right circular truncated cones 4, 6 medium cylinders 5 and 12 elastic sheets 6. Wherein, the disc type centrifugal bracket 1 is connected with a nucleic acid detector.
The central circular truncated cone 3 is positioned at the central position of the upper surface of the disc type centrifugal support 1, and the bottom surface of the central circular truncated cone 3 faces downwards.
The small-sized right circular truncated cone 4 abuts against the central right circular truncated cone 3 and is uniformly distributed, the size of the cone angle is equal to that of the cone angle of the central right circular truncated cone 3, the bottom surface of the small-sized right circular truncated cone 4 faces upwards, and the small-sized right circular truncated cone has downward friction force, so that the small-sized right circular truncated cone has a buckling effect and buckles the clamping groove 7 on the small arc edge of the fan-shaped.
The medium-sized cylinders 5 are uniformly distributed on the upper surface of the disc type centrifugal support 1 and close to the edge of the disc type centrifugal support 1, each medium-sized cylinder 5 corresponds to the middle position of each 2 small circular truncated cones 4, and each 2 medium-sized cylinders 5 fix 1 fan-shaped chip 2.
The elastic pieces 6 are uniformly distributed on the edge of the disc type centrifugal support 1, the positions of the elastic pieces are arranged at a distance of about 5 mm protruding out of the disc type centrifugal support 1, the middle position of each 2 elastic pieces 6 corresponds to the small right circular table 4, the angle a of each elastic piece 6 is less than 80 degrees, and the elastic pieces are used for fixing 1 fan-shaped chip 2 and play a role in buffering.
The fan-shaped chips 2 are made of transparent materials, the transparent materials comprise PMMA, the vertex angle of each fan-shaped chip 2 is 45 degrees, the fan-shaped chips 2 are fixed among every 2 medium-sized cylinders 5, and the reaction holes 11 are exposed on the bottom plate of the disc type centrifugal support 1.
The fan-shaped chip 2 consists of a clamping groove 7, a liquid adding pool 8, a liquid separating groove 9, a gas replacement hole 10, a reaction hole 11 and a waste liquid groove 12 which are connected through a flow passage 13.
The clamping groove 7 is a half small right circular truncated cone and is positioned at the center of the small arc edge of the fan-shaped chip 2 and just matched with the small right circular truncated cone 4, and the small right circular truncated cone 4 is fixed due to downward friction force.
The volume of the liquid adding pool 8 is larger than 60 mu l, the end far away from the circle center is connected with the liquid separating groove 9 through the flow channel 13, a sample larger than 60 mu l is firstly added into the liquid adding pool 8, the sample flows into the 5 cup-shaped liquid separating grooves 15 of the liquid separating groove 9 under the action of centrifugal force, and after the liquid separating groove 9 collects the sample, the gas is displaced through the gas displacement holes 10 and the reaction holes 11, so that the 5 reaction holes 11 are filled with the sample, and the redundant sample can be discharged into the waste liquid groove 12.
The sealing adhesive film 14 is transparent, completely covers and adheres to the surface of the fan-shaped chip 2, a sample adding hole 16 is formed in the position, far away from the circle center end, corresponding to the sample adding pool 8, the diameter of the sample adding hole 16 is 1mm, and a sample is added into the sample adding pool 8 from the sample adding hole 16.
The fan-shaped chip 2 realizes the equal-amount split charging of liquid through centrifugal force and the reaction hole 11.
Different reagents, primers or probes can be pre-buried in the 5 reaction holes 11, and reaction detection is synchronously performed through sample adding and centrifugation.
The sealing film 14 contains a removable sample hole film 17, and the sample hole film 17 just covers the sample hole 16.
The multi-joint detection device for the sector nucleic acid consists of three components, namely 1 disc type centrifugal support 1, 6 sector chips 2 and 6 closed adhesive films 14. The disc-type centrifugal support 1 can be installed in a nucleic acid detector, and the upper surface of the disc-type centrifugal support is provided with 6 sectors A-F corresponding to 1 central right circular truncated cone 3, 6 concentric small right circular truncated cones 4, 6 medium cylinders 5 and 12 elastic sheets 6. The central right circular truncated cone 3 is positioned at the central position of the upper surface of the disc type centrifugal bracket 1, and the bottom surface of the circular truncated cone faces downwards. The small-sized right circular truncated cone 4 abuts against the central right circular truncated cone 3 and is uniformly distributed, the size of the cone angle is equal to that of the cone angle of the central right circular truncated cone 3, the bottom surface of the circular truncated cone faces upwards, and the circular truncated cone has downward friction force, so that the clamping effect is realized, and the clamping groove 7 on the small arc edge of the fan-shaped chip 2 is buckled. The medium-sized cylinders 5 are uniformly distributed on the upper surface of the support and close to the edge of the support, each medium-sized cylinder 5 corresponds to the middle position of each 2 small-sized right circular truncated cones 4, and each 2 medium-sized cylinders 5 fix 1 fan-shaped chip 2. The elastic sheets 6 are uniformly distributed on the edge of the disc type centrifugal support 1, protrude out of the disc type centrifugal support 1 by about 5 mm, the middle position of each 2 elastic sheets 6 corresponds to the small-sized right circular table 4, the angle a of each elastic sheet 6 is less than 80 degrees, and the elastic sheets are used for fixing 1 fan-shaped chip 2 and play a role in buffering.
The fan-shaped chips 2 are made of transparent materials, the transparent materials comprise PMMA and the like, the vertex angle of each fan-shaped chip 2 is 45 degrees, the fan-shaped chips are fixed among every 2 medium-sized cylinders 5, every small-sized circular truncated cone 4 and 2 elastic sheets 6, and the reaction holes 11 are exposed on the bottom plate of the disc type centrifugal support 1. The fan-shaped chip 2 is composed of a clamping groove 7, a liquid adding pool 8, a liquid separating groove 9, a gas replacement hole 10, a reaction hole 11 and a waste liquid groove 12, wherein the clamping groove 7, the liquid adding pool 8, the liquid separating groove 9, the gas replacement hole 10, the reaction hole 11 and the waste liquid groove 12 are connected through a flow channel 13. The clamping groove 7 is a half small right circular truncated cone and is positioned at the center of the small arc edge of the fan-shaped chip 2 and just matched with the small right circular truncated cone 4, and the small right circular truncated cone 4 is fixed due to downward friction force. The volume of the liquid adding pool 8 is more than 60 mul, and the end far away from the circle center is connected with the liquid separating groove 9 through a flow passage 13. More than 60 μ l of sample is first added to the addition cell 8, and the sample flows into the 5-cup type separation cell 15 of the separation cell 9 by the centrifugal force, and after the collection of the sample in the separation cell 9 is completed, the gas is displaced through the gas displacement holes 10 and the reaction holes 11, so that the 5 reaction holes 11 are filled with the sample, and the excess sample is discharged into the waste liquid cell 12.
The sealing pad pasting 14 is transparent, completely covers and pastes on the surface of the fan-shaped chip 2, a position corresponding to the end of the sample adding pool 8 far away from the center of the circle contains a sample adding hole 16 and a pad pasting 17 covering the sample adding hole, the diameter of the sample adding hole 16 is 1mm, and a sample is added into the sample adding pool 8 from the sample adding hole 16.
After the sector chip 2 is balanced, the equivalent split charging of the liquid is realized through centrifugal force. Different reagents, primers or probes can be pre-buried in the 5 reaction holes 11, and fluorescence detection is synchronously performed through sample adding, balancing, centrifuging and heating.
In specific implementation, reagents, primers or probes are pre-buried in the reaction holes 11, a dry powder is formed after freeze-drying, and the sealing adhesive film 14 is attached to form the chip for nucleic acid detection. During detection, 60 mu l of processed sample is taken by a pipette and slowly added into a sample adding pool 8 from a sample adding hole 16, a pad pasting is pasted to cover the sample adding hole, a chip is installed on a disc type centrifugal support 1 in a nucleic acid detector, after balancing, the sample is evenly subpackaged and filled in the whole reaction hole 11 through the centrifugation of a machine, the machine is heated at 37-45 ℃ for constant temperature, each reaction hole 11 completes the nucleic acid amplification of corresponding indexes, the machine collects the fluorescence signals of each reaction hole 11, data processing and graph drawing are carried out, the result is automatically judged and read, and the detection of multiple indexes of a single sample is realized.
The utility model relates to a fan-shaped many joint inspection of nucleic acid device, each reaction hole 11 of the device can be pre-buried reagent, primer or probe etc. of different projects respectively, combines EMA technique, can realize sample single sample, 37-45 ℃ constant temperature reaction, obtains many index testing results in 30 minutes. The utility model has the advantages of sample reagent low consumption, easy operation, the leakproofness is good, pollutes for a short time, high flux, low cost practices thrift patient check-out time and cost simultaneously, is fit for being applied to the examination of clinical pathogen, assists scientific medication.
Example 1 simulation experiment for use of chip
As shown in FIGS. 1 and 2, a nucleic acid detecting chip is formed by embedding water (< 5. mu.l) into the reaction well 11 and attaching a sealing film 14 thereto. In the detection, the well pad 17 is uncovered, 60 μ l of the simulated sample is pipetted by a pipette, slowly added into the sample cell 8 from the well 16, and the well pad 17 is attached again to cover the well 16. The sector chip 2 is arranged on a disc type centrifugal support 1 in a nucleic acid detector, balancing is carried out, a sample is uniformly subpackaged and filled in the whole reaction hole 11 through the centrifugation of a machine, the machine is heated at the constant temperature of 37-45 ℃, the fluorescence signals of all holes are collected by the machine, and data processing and graph drawing are carried out.
EXAMPLE 2 verification of Vent hole
As shown in FIG. 4, the waste liquid tank 12 is not connected to the liquid addition tank 8, and a small hole is pierced at a position corresponding to the air discharge hole 18 of the well patch 17 for air discharge when in use. The sample is loaded into the sample loading well 8 and the loading well 16 is closed as in example 1. The small hole in the vent hole 18 is not closable because of the need to vent the gas. The sector chip 2 is installed on a disc type centrifugal support 1 in the nucleic acid detector, the sector chip 2 shown in figure 2 is used as a contrast, the sector chip 2 is taken out after the operation is finished through the centrifugation of a machine, the reaction holes 11 of the two sector chips 2 are both bubble-free, but considering that the exhaust holes 18 on the sector chip 2 are not closed, the risk of aerosol pollution is generated, and the risk of polluting the detection environment is increased. The waste liquid tank 12 is communicated with the addition tank 8.
EXAMPLE 3 preparation and testing of respiratory tract 5 Joint inspection chip
As shown in FIG. 2, in the reaction wells 11, the reagents of each reaction well 11 pass through the microfluidic pre-embedded reagent, the reagent concentration of each reaction well 11 comprises a single-stranded DNA binding protein with a final concentration of 24-27ng/μ l, an ATP regeneration protein with a final concentration of 62-125pg/μ l, a DNA helicase with a final concentration of 6-7.5ng/μ l, a DNA polymerase with a final concentration of 2-3ng/μ l, a DNA restriction endonuclease with a final concentration of 90-120pg/μ l, an accessory protein with a final concentration of 300 pg/μ l, creatine phosphate sodium with a concentration of 20-30mM, ATP with a final concentration of 2-3mM, dNTP with a concentration of 100-150 μ M, Tris-Ac with a concentration of 50-60mM (pH8.0), trehalose with a final concentration of 35-50 μ g/μ l, KAc with a concentration of 100-120mM, mannitol with a final concentration of 7.5-10ng/μ l, PEG20000(W/V), wherein the RNA system is additionally added with a final concentration of 0.75-7.5ng/μ l M-, 0.25-2.5 ng/. mu.l RNase inhibitor.
5 reaction holes 11 are pre-embedded with 5 items of primers and probes respectively, clockwise sequentially comprise mycoplasma pneumoniae, chlamydia pneumoniae, syncytial virus, influenza virus H3 subtype and legionella pneumophila, the concentration of the primers is 200-300nM, the concentration of the probes is 30-40nM, and the sequences are shown in Table 1. After the reagents are pre-buried, freeze-drying is carried out, and a sealing film is attached to form a simple multi-detection chip.
TABLE 1 primer and probe sequences for respiratory tract 5 joint inspection of each reaction well
Note: "i 6-FAMdT" refers to 6-FAM (6-carboxyfluorescein) fluorescently labeled dT nucleotide, "idSp" refers to base deletion, "iBHQ 1 dT" refers to dT nucleotide labeled with BHQ1 quencher group, and "C3 Spacer" refers to 3' hydroxyl blocking.
For detection, the mock sample used in this example was an equal mixture containing plasmids of the target sequences of Mycoplasma pneumoniae, Chlamydia pneumoniae, and Legionella pneumophila. And (3) adding 30 mu l of the simulated sample into 30 mu l of the reaction solution by using a liquid transfer gun, sucking and uniformly mixing the mixture, wherein the reaction solution contains MgAc2 with the final concentration of 20-100mM, Tris-Ac with the final concentration of 200mM, KAc with the final concentration of 140mM of 110-100 mM and PEG20000 with the final concentration of 10-15%, uncovering the sample adding hole adhesive film 17, transferring the sample nucleic acid into the sample adding pool 8 from the sample adding hole 16 by using the liquid transfer gun, attaching the sample adding hole adhesive film 17 again, and sealing the sample adding hole 16. And (3) installing the sector chip 2 after sample adding into a sector A of the disc type centrifugal support 1, balancing, clockwise marking 1-5 reaction holes in the area A, namely A1-A5, and respectively detecting whether the sample contains mycoplasma pneumoniae, chlamydia pneumoniae, syncytial virus, influenza virus H3 subtype or legionella pneumophila.
Selecting a reaction program on a machine and operating, centrifuging at 5000rpm for 1min, uniformly subpackaging and filling the samples in the whole reaction holes 11, heating the machine at the constant temperature of 37-45 ℃, completing nucleic acid amplification of corresponding indexes in each reaction hole, collecting fluorescence signals of each hole at the interval of 30s by the machine, processing data and drawing graphs to give a report, wherein the result is shown in figure 5. Three holes A1, A2 and A5 show 3 curves, which are positive results of mycoplasma pneumoniae, chlamydia pneumoniae and legionella pneumophila respectively, while the amplification curves of A3 and A4 are not shown, and the syncytial virus and influenza virus H3 subtypes show negative results, which indicates that the sample contains the target sequences of genes of the mycoplasma pneumoniae, the chlamydia pneumoniae and the legionella pneumophila and is consistent with the expected results.
The utility model discloses a fan-shaped many joint inspection devices of nucleic acid comprises disc type centrifugal support, fan-shaped chip and closed pad pasting. The device can realize the simultaneous amplification and detection of multiple projects of single sample, and is higher in efficiency than a common nucleic acid detection device, good in sealing performance and capable of effectively avoiding product aerosol pollution. Meanwhile, the consumption of samples, consumables and reagents is reduced, the pathogen omission probability is reduced, the detection flux is improved, and the kit is suitable for being applied to clinical pathogen screening and assisting in scientific medication.
The fan-shaped nucleic acid multi-joint detection device has the advantages that:
(1) the utility model provides a fan-shaped nucleic acid multi-joint inspection device, can realize single sample and obtain many index testing results, realize high flux screening, practiced thrift patient's detection cost and time;
(2) the utility model provides a fan-shaped nucleic acid multi-detection device, the operation process of detection personnel is simplified, only two steps of sample adding and film pasting are needed, the error rate is reduced, and the personnel cost and time are saved;
(3) the utility model provides a fan-shaped nucleic acid multi-joint inspection device, reagent and the like are all filled in advance, thereby reducing the exposure of the reagent in the operation process of the inspector and reducing the probability of false positive;
(4) the utility model provides a fan-shaped nucleic acid multi-detection device, after the application of sample is accomplished, a closed space is formed by the covering of the pad pasting, which avoids the evaporation of aerosol and reduces the pollution;
(5) the utility model provides a fan-shaped nucleic acid multi-detection device, the waste liquid groove is communicated with the sample adding groove, which ensures the smooth liquid flow under the action of centrifugal force;
(6) the utility model provides a fan-shaped nucleic acid multi-joint inspection device, the arrangement of the gas replacement hole can ensure that the whole reaction hole can be filled with the reagent, and the reaction accuracy is improved;
(7) the utility model provides a fan-shaped nucleic acid multi-link inspects device, the device haplopore reaction volume is 10 ul, has reduced the consumption of sample, consumptive material and reagent, has practiced thrift manufacturing cost.
The above is only the preferred embodiment of the present invention, and it should be noted that, for those skilled in the art, many variations and modifications can be made without departing from the inventive concept, and these all fall into the protection scope of the present invention.
Claims (9)
1. A fan-shaped nucleic acid multi-joint detection device is characterized by comprising: disc type centrifugal support, fan-shaped chip and closed pad pasting, disc type centrifugal support with fan-shaped chip is connected, fan-shaped chip with closed pad pasting is connected, fan-shaped chip includes: the device comprises a liquid adding pool, a liquid separating tank, a gas displacement hole, a reaction hole and a waste liquid tank, wherein the liquid adding pool, the liquid separating tank, the gas displacement hole, the reaction hole and the waste liquid tank are connected with one another through flow channels.
2. The fan-shaped nucleic acid multi-gang detection device of claim 1, wherein a central circular truncated cone is arranged on the disc-type centrifugal support, and the central circular truncated cone is connected with the disc-type centrifugal support.
3. The fan-shaped nucleic acid multi-detection device according to claim 2, wherein a small positive circular table is arranged on the disc-type centrifugal support and connected with the disc-type centrifugal support, and the small positive circular table is uniformly arranged along the circumference of the central positive circular table and clamped with a clamping groove on one side of the fan-shaped chip.
4. The fan-shaped nucleic acid multi-joint inspection device according to claim 3, wherein a spring plate is arranged on the disc-shaped centrifugal support, and the other side of the fan-shaped chip is fixedly connected with the disc-shaped centrifugal support through the spring plate.
5. The multi-gang nucleic acid analyzer as claimed in claim 1, wherein the sample is added to the liquid feeding well, the sample flows into the liquid separating tank by the centrifugal force of the disc-type centrifugal support, the liquid separating tank collects the sample, and the gas is displaced through the gas displacement holes and the reaction holes, so that the reaction holes are filled with the sample, and the excess sample is discharged to the waste liquid tank.
6. The fan-shaped multi-unit nucleic acid detection device according to claim 5, wherein the sealing film is provided with sample adding holes corresponding to and cooperating with the liquid adding pool for adding the sample into the liquid adding pool from the sample adding holes.
7. The fan-shaped nucleic acid multi-detection device according to claim 6, wherein the sample adding holes are provided with sample adding hole adhesive films, and the sample adding hole adhesive films correspond to the sample adding holes and are connected with the sample adding holes in a matched manner.
8. The multiple nucleic acid testing device as claimed in claim 7, wherein six groups of medium-sized cylinders are arranged on the disc-type centrifugal rack.
9. The fan-shaped nucleic acid multi-gang detection device of claim 8, wherein six groups of medium-sized cylinders are uniformly distributed on the edge of the surface of the disc-type centrifugal rack, and the fan-shaped chips are arranged between and fixedly connected with the two groups of medium-sized cylinders.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201920387799.2U CN210394313U (en) | 2019-03-28 | 2019-03-28 | Fan-shaped nucleic acid multi-joint detection device |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109810875A (en) * | 2019-03-28 | 2019-05-28 | 苏州点晶生物科技有限公司 | A kind of multi-joint checking device of sector nucleic acid |
| WO2023173640A1 (en) * | 2022-03-16 | 2023-09-21 | 广州达安基因股份有限公司 | Cartridge for nucleic acid extraction and quantitative liquid separation |
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2019
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109810875A (en) * | 2019-03-28 | 2019-05-28 | 苏州点晶生物科技有限公司 | A kind of multi-joint checking device of sector nucleic acid |
| WO2023173640A1 (en) * | 2022-03-16 | 2023-09-21 | 广州达安基因股份有限公司 | Cartridge for nucleic acid extraction and quantitative liquid separation |
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