CN109439737A - A kind of kit and its diagnostic device for cerebrospinal fluid difficulty pathogen diagnosis - Google Patents
A kind of kit and its diagnostic device for cerebrospinal fluid difficulty pathogen diagnosis Download PDFInfo
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- CN109439737A CN109439737A CN201910018294.3A CN201910018294A CN109439737A CN 109439737 A CN109439737 A CN 109439737A CN 201910018294 A CN201910018294 A CN 201910018294A CN 109439737 A CN109439737 A CN 109439737A
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Classifications
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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
- C12Q1/6876—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
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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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- 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
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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
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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
- C12Q2600/00—Oligonucleotides characterized by their use
- C12Q2600/16—Primer sets for multiplex assays
Abstract
This application discloses a kind of kits for cerebrospinal fluid difficulty pathogen diagnosis, not only include reagent box body, further include fixed plate, primer plate and 96 hole PCR reaction plates positioned at reagent box body；The first groove and the second groove are provided in fixed plate, the number of the second groove and the number of centrifuge tube are corresponding；Be provided on primer plate can drop target primer multiple test tubes third groove, the number of third groove and the number of test tube are corresponding, and test tube is equipped with a kind of target primer, there is the lid of helixseal on test tube.It include the primer plate for holding multiple test tubes of plurality of target primer in the kit, actually detected how many kinds of primer, how many a third grooves are just set on primer plate, place how many a test tubes, realize the diagnosis in multi-infection source, it can ensure that clinical precisely diagnosis pathogen infection type and adjustment pesticide application strategy, reduce the death rate.In addition, disclosed herein as well is a kind of cerebrospinal fluid difficulty pathogen diagnostic device, effect is as above.
Description
Technical field
This application involves medical field more particularly to a kind of kit for cerebrospinal fluid difficulty pathogen diagnosis and its examine
Disconnected device.
Background technique
Currently, very nervous centralis infection, encephalitis, most of meningitis are all as caused by various pathogen, therefore, to each
The quick detection of kind pathogen is particularly important.Not so, higher case fatality rate, hospitalization cost rising and bed turnover will be caused
The health economics problems such as number of days increase.
Now, the main detection realized by diagnostic kit to the cerebrospinal fluid source of infection, still, traditional detection mode is only
Can judge whether patient infects a kind of special pathogen, and can not determine patient whether the cause of disease also infected with other types
Body, detectable infection source category is few, and diagnostic accuracy is lower, and then prevents diagnosis person from clinically accurate medication, can be wrong
The golden hour of patient is spent, What is more will lead to patient and fortuitous event occur.
It can be seen that how increasing the detection type of the source of infection, improving the detection accuracy of the source of infection so that clinical accurate
The problem of diagnosing pathogen infection type and adjustment pesticide application strategy, reducing mortality is that those skilled in the art are urgently to be resolved
The problem of.
Summary of the invention
This application provides a kind of kit and its diagnostic device for cerebrospinal fluid difficulty pathogen diagnosis, solve existing
There is the detection type for how increasing the source of infection in technology, improve the detection accuracy of the source of infection so that clinical precisely diagnosis infection disease
The problem of substance type and adjustment pesticide application strategy, reduction mortality.
In order to solve the above technical problems, this application provides a kind of kit for cerebrospinal fluid difficulty pathogen diagnosis,
Including reagent box body, further includes:
Positioned at the fixed plate of the reagent box body, primer plate and 96 hole PCR reaction plates；
The first groove for fixing the primer plate and for placing each centrifuge tube are provided in the fixed plate
Two grooves, the number of second groove and the number of the centrifuge tube correspond；
It is provided with the third groove for placing multiple test tubes that plurality of target primer is housed on the primer plate, described the
The number of three grooves and the number of the test tube correspond, and a test tube is equipped with a kind of target primer, the examination
Pipe exit is equipped with the lid of helixseal.
Preferably, the sealing ring for sealing is additionally provided on the lid of the helixseal.
Preferably, each second groove is located at the same side of the fixed plate.
Preferably, each second groove is uniformly distributed in the same side of the fixed plate.
Preferably, the cross section of the primer plate is rectangle；
Accordingly, the cross section of first groove is rectangle.
Preferably, further includes:
It is installed on the primer plate, the lid being engaged positioned at the edge of third groove side and the primer plate.
In order to solve the above technical problems, present invention also provides a kind of reagents with for cerebrospinal fluid difficulty pathogen diagnosis
The corresponding cerebrospinal fluid difficulty pathogen diagnostic device of box is used for the examination of cerebrospinal fluid difficulty pathogen diagnosis based on any one of the above
Agent box, comprising:
First obtains module, utilizes centrifuge tube storage in kit for obtaining according to the first acquisition instruction received
The DNA information extracted from cerebrospinal fluid；
Second obtains module, for obtaining a variety of primers according to the second acquisition instruction received；
First determining module, for determining target primer present in DNA according to the DNA information；
Second determining module, for determining corresponding antibiotic in the cerebrospinal fluid according to the target primer
Pathogen is diagnosed；
Wherein, the primer includes the target primer.
Preferably, first determining module is specifically used for determining the target primer by PCR-Array mode.
Preferably, first determining module specifically includes:
Judgment module, for judging whether each primer is positive in the DNA, if it is, triggering third is true
Cover half block, if it is not, then the 4th determining module of triggering；
Third determining module, for determining, there are the primers in the DNA, and the primer is known as the target and is drawn
Object；
4th determining module, for determining, there is no the primers in the DNA.
Preferably, the type of the primer is specially 20 kinds, respectively escherichia coli, enterococcus faecalis, streptococcus pneumonia,
Serratia marcescens, proteus mirabilis, stenotrophomonas maltophilia, Acinetobacter bauamnnii, pseudomonas aeruginosa, dung intestines ball
Bacterium, klebsiella pneumoniae, staphylococcus aureus, Bu Shi citric acid bacillus, Burkholderia cepacia, epidermis grape ball
Bacterium, Candida albicans, Candida tropicalis, Candida parapsilosis bacterium, Candida glabrata, candida krusei, aspergillus fumigatus, list
Pure bleb, HHV-6, CMV and VZV.
Compared with the prior art, a kind of kit for cerebrospinal fluid difficulty pathogen diagnosis provided herein, is removed
Include except reagent box body, further includes fixed plate, primer plate and the 96 hole PCR reaction plates positioned at reagent box body；Gu
It is provided on fixed board for the first groove of immobilized primer plate and the second groove for placing each centrifuge tube, of the second groove
Several numbers with centrifuge tube correspond；The for placing multiple test tubes that plurality of target primer is housed is provided on primer plate
Three grooves, the number of third groove and the number of test tube correspond, and a test tube is equipped with a kind of target primer, test tube exit
The lid of helixseal is installed.That is, including multiple test tubes for holding plurality of target primer in the kit
Primer plate, actual needs detection how many kinds of primer (source of infection) is just arranged how many a third grooves on primer plate, and placement is more
Few test tube, may be implemented the checkout and diagnosis to multi-infection source, improve the detection accuracy of the source of infection, and then can clinical essence
Quasi- diagnosis pathogen infection type and adjustment pesticide application strategy reduce mortality and shorten patient because being hospitalized week caused by infection
Phase.In addition, present invention also provides a kind of cerebrospinal fluid difficulty pathogen diagnostic device, effect is as above.
Detailed description of the invention
For the clearer technical solution for illustrating the application, letter will be made to attached drawing needed in the embodiment below
The introduction wanted, it should be apparent that, it for those of ordinary skills, can be under the premise of not paying creativeness
It obtains other drawings based on these drawings.
Fig. 1 is a kind of reagent cartridge configuration figure for cerebrospinal fluid difficulty pathogen diagnosis provided by the embodiment of the present invention；
Fig. 2 is a kind of cerebrospinal fluid difficulty pathogen diagnostic device composition schematic diagram provided by the embodiment of the present invention.
Specific embodiment
In order to make those skilled in the art better understand the technical solutions in the application, below in conjunction with attached drawing, it is right
Technical solution in the embodiment of the present application carries out clear and complete description.
The core of the application is to provide a kind of kit and its diagnostic device for cerebrospinal fluid difficulty pathogen diagnosis, can
To solve how to increase in the prior art the detection type of the source of infection, improve the detection accuracy of the source of infection so that clinical precisely examine
The problem of disconnected pathogen infection type and adjustment pesticide application strategy, reduction mortality.
Fig. 1 is a kind of reagent cartridge configuration figure for cerebrospinal fluid difficulty pathogen diagnosis provided by the embodiment of the present invention,
As shown in Figure 1, the diagnostic kit includes reagent box body 1, further includes:
Positioned at the fixed plate 10 of reagent box body 1, primer plate 11 and 96 hole PCR reaction plates 12；
It is provided in fixed plate 10 for the first groove 100 of immobilized primer plate 11 and for placing each centrifuge tube 13
Two grooves 101, the number of the second groove 101 and the number of centrifuge tube 13 correspond；
The third groove 110 for placing multiple test tubes 14 that plurality of target primer is housed is provided on primer plate 11, the
The number of three grooves 110 and the number of test tube 14 correspond, and a test tube 14 is equipped with a kind of target primer, 14 exit of test tube
The lid 140 of helixseal is installed.
Specifically, reagent box body 1 mainly just refers to the shell of kit, can be carton box, is also possible to plastic casing.
In the embodiment of the present application, the fixed plate 10 inside kit can select cystosepiment, be provided in fixed plate 10 for placing
First groove 100 of primer plate 11, can make primer plate 11, completely or partially fixation is embedded in first to the size of the first groove 100
Subject to groove 100.The second groove 101 for placing each centrifuge tube 13 (EP pipe), centrifuge tube 13 are additionally provided in fixed plate 10
It is mainly used for holding Cerebrospinal Fluid in Patients, the reaction solution that the DNA extracting solution that extracts from the cerebrospinal fluid and when experiment need；In reality
In the application of border, related personnel obscures the substance that each centrifuge tube 13 is filled in order to prevent, and then influences diagnostic result, can also be each
Etch layer is added on centrifuge tube 13, so as to the printed words of the engraving " this centrifuge tube is equipped with XX substance " in the etch layer, is examined with improving
Disconnected accuracy；The number of second groove 101 and the number of centrifuge tube 13 correspond, that is, have several centrifuge tubes 13, just exist
Several second grooves 101 are set in fixed plate 10, and the number of centrifuge tube 13 can be determined according to the actual situation, the second groove
101 size, which is subject to, can make all or part of fixation of each centrifuge tube 13 be embedded in the second groove 101；In view of the convenience of design
The aesthetics of property and finished product, preferably embodiment, each second groove 101 are located at the same side of fixed plate 10, simultaneously
Ensure that each second groove 101 is uniformly distributed in the same side of fixed plate 10, i.e., the same left or right side for being uniformly located at fixed plate 10,
When the first groove 100 is located at the left side of fixed plate 10, each second groove 101 is uniformly located at the right side of fixed plate 10, when first
When groove 100 is located at the right side of fixed plate 10, each second groove 101 is uniformly located at the left side of fixed plate 10, and certainly, each second is recessed
Slot 101 can also be distributed in other ways in fixed plate 10, it is not limited to the distribution mode in the embodiment of the present application.In reality
In the application of border, the size of the kit in the embodiment of the present application can be determined according to the actual situation, and the present invention is simultaneously not construed as limiting.
Primer plate 11 inside kit can select rubber slab, be provided on primer plate 11 for placing multiple test tubes 14
Third groove 110, the number of test tube 14 needs the type of target primer according to actual needs to determine, a test tube 14 is equipped with one
Kind of target primer, for example, if actual needs 29 kinds of primers of diagnosis, it is necessary to 29 test tubes 14, primer be also referred to as pathogen or
The source of infection, the number of third groove 110 and the number of test tube 14 correspond, that is to say, that have several test tubes 14, it is necessary to
Several third grooves 110 are set on primer plate 11, and the size of third groove 110 is can make all or part of fixed edge of each test tube 14
Subject to third groove 110, in actual design, third groove 110 can be made to be uniformly distributed in primer plate 11；Actually answering
In, the primer filled in each test tube 14 is accurately quickly identified for the ease of related personnel, improves diagnosis efficiency, it can also be each
Etch layer is added on test tube 14, so as to the printed words of the engraving " primer filled in this test tube 14 is XX " in the etch layer, to improve
Diagnosis efficiency；In view of the characteristic of each target primer, 14 exit of test tube need to install the lid 140 of helixseal, helixseal
The material of lid 140 can be determined according to the actual situation.In view of the face shaping of centrifuge tube 13 and test tube 14, in reality
When setting, the cross section of the second groove 101 and third groove 110 can be disposed as circle, certainly, in practical applications,
The cross section of second groove 101 and third groove 110 can also be other shapes, as long as may insure 13 whole of centrifuge tube or portion
It point is capable of fixing the second groove 101 of insertion and test tube 14 is all or part of is capable of fixing insertion third groove 110.It considers
The convenience of design, preferably embodiment is in the cross section of primer plate 11 is rectangle；Accordingly, the first groove 100
Cross section be rectangle, particularly as be primer plate 11 cross section be which kind of shape, the cross section of the first groove 100 is just designed
For which kind of shape, i.e. saving material, it is also convenient for immobilized primer plate 11.It, can be by fixed plate in view of the convenience of practical operation
10 and 96 hole PCR reaction plates 12 are laid in the bottom position of reagent box body 1, it is, of course, also possible to by the hole of fixed plate 10 and 96
PCR reaction plate 12, which stacks, to be placed, and the present invention is simultaneously not construed as limiting.It, can be in view of the rapidity of design and the aesthetics of appearance
Reagent box body 1 is made into square.Certainly, the concrete shape of reagent box body 1 is not limited in the embodiment of the present application
Shape can also be designed to that other shapes, the present invention are simultaneously not construed as limiting.
It is typically provided with 96 reactive tanks 120 on 96 hole PCR reaction plates 12, is used to store reaction solution in reactive tank 120
(mixture of DNA extracting solution and plurality of target primer), the structure of 96 hole PCR reaction plates 12 and 96 hole PCR in the prior art
The structure of reaction plate is the same, and for details, reference can be made to the prior arts, and details are not described herein by the present invention.
The condition of storage that the application implements provided kit is to be kept in dark place, and is avoided pollution, -20 DEG C of transports；-20
DEG C long-term preservation；Multigelation does not influence using effect；If frequently using, it is proposed that be stored in 4 DEG C, 4 DEG C can be reserved for 6 months.If
Part solution generates precipitating, the solution in kit should be placed at room temperature for a period of time before use, when necessary can be in 37 DEG C of water
10min is preheated in bath, to dissolve precipitating.Points for attention are to need to read over this kit specification before experiment, in strict accordance with saying
Bright book executes operation.It should illustrate that anhydrous second is added in elder generation in buffer and rinsing liquid according to reagent bottle label before first time use
Alcohol.Multigelation should be avoided in sample, and the DNA fragmentation that otherwise will lead to extraction is smaller and extracted amount also declines.If buffer is clear
There is precipitating in washing lotion, can re-dissolve in 37 DEG C of water-baths, be used after shaking up.All centrifugation steps are to use desk-top centrifugation
Machine is centrifuged at room temperature.Sample should be balanced before bringing into operation to room temperature (15~25 DEG C).It is best to long-term preservation DNA sample
The dehydrated alcohol of corresponding 2.5 times of volumes is added to precipitate DNA.White Flocculus can be seen after mixing.It is placed after of short duration centrifugation
At least a year is saved in -80 DEG C of low temperature refrigerators.If needing using need to be centrifuged 5min in the centrifuge of 7000~8000 × g, make
DNA is precipitated again, adds TE solution dissolving DNA precipitating.The purity and concentration of DNA must be detected again when necessary.MasterMix
Should not multigelation, if be commonly used, preferably dissolve after be placed on 4 DEG C.After frozen product dissolution, it please turn upside down gently uniformly
Mixing, is sure not excessively to exert oneself, avoids blistering, and use after gentle centrifugation.Reagent after melting please is placed on ice.Reaction is matched
Set, dispense (free of contamination) pipette tips (be not loaded with the same pipette tips continuously), the miniature tube for having to renew etc., it avoids as far as possible
Pollution.Gloves will be worn during using the kit, if gloves are contacted with sample, need to replace gloves immediately.
A kind of kit for cerebrospinal fluid difficulty pathogen diagnosis provided herein, in addition to including reagent box body
Except, it further include fixed plate, primer plate and the 96 hole PCR reaction plates positioned at reagent box body；It is provided with and is used in fixed plate
First groove of immobilized primer plate and the second groove for placing each centrifuge tube, the number of the second groove and the number of centrifuge tube
It corresponds；The third groove for placing multiple test tubes that plurality of target primer is housed, third groove are provided on primer plate
Number and the number of test tube correspond, test tube is equipped with a kind of target primer, and test tube exit is equipped with helixseal
Lid.That is, include the primer plate for holding multiple test tubes of plurality of target primer in the kit, it is practical to need
How many kinds of primer (source of infection) is detected, how many a third grooves are just set on primer plate, places how many a test tubes, Ke Yishi
Now to the checkout and diagnosis in multi-infection source, the detection accuracy of the source of infection is improved, and then may insure clinical precisely diagnosis infection
Pathogen type and adjustment pesticide application strategy reduce mortality and shorten patient because being hospitalized the period caused by infection.
On the basis of Fig. 1, the kit further include: be installed on primer plate 11, be located at 110 side of third groove and primer
The lid that the edge of plate 11 is engaged.Particularly as being to be additionally provided with lid on primer plate 11, lid can be with the edge of primer plate 11
Be interlocked, in practical applications, can by lid it is any on one side be fixedly installed in primer plate 11 while position place (because drawing
The cross section of object plate 11 is rectangle, so the cross section of matched lid also should be rectangle), remaining other three
While closely being fastened when lid is detained with the edge of primer plate 11, it can prevent test tube 14 in primer plate 11 and air long
Time contact, can also shake in the third groove 110 on primer plate 11 to avoid test tube 14, if lid is light screening material,
It can also play the role of being protected from light.In order to further prevent omit air enter in test tube 14, influence diagnosis accuracy,
On the basis of above-described embodiment, preferably embodiment is additionally provided with for the close of sealing on the lid 140 of helixseal
Seal.Particularly as being to add one layer of sealing ring again on the lid 140 of helixseal.
It is described in detail above for a kind of embodiment of kit for cerebrospinal fluid difficulty pathogen diagnosis,
The kit for cerebrospinal fluid difficulty pathogen diagnosis described based on the above embodiment, the embodiment of the invention also provides one kind
Cerebrospinal fluid difficulty pathogen diagnostic device corresponding with the kit.Due to the embodiment of device part and the reality of kit of parts
Example reciprocal correspondence is applied, therefore the embodiment of device part please refers to the embodiment description of kit of parts, which is not described herein again.
Fig. 2 is a kind of cerebrospinal fluid difficulty pathogen diagnostic device composition schematic diagram provided by the embodiment of the present invention, such as Fig. 2
It is shown, based on the kit for being used for cerebrospinal fluid difficulty pathogen diagnosis provided by any one above-mentioned embodiment, diagnosis dress
It sets and obtains module 201 including first, second obtains module 202, the first determining module 203 and the second determining module 204.
First obtains module 201, is deposited for being obtained according to the first acquisition instruction received using centrifuge tube in kit
The DNA information extracted in the slave cerebrospinal fluid put；
Second obtains module 202, for obtaining a variety of primers according to the second acquisition instruction received；
First determining module 203, for determining target primer present in DNA according to DNA information；
Second determining module 204, for determining corresponding antibiotic to the cause of disease in cerebrospinal fluid according to target primer
Body is diagnosed；
Wherein, primer includes target primer.
On the basis of the above embodiments, preferably embodiment, the first determining module 201 are specifically used for passing through
PCR-Array mode determines target primer.
On the basis of the above embodiments, preferably embodiment, the first determining module 201 specifically include:
Judgment module, for judging whether each primer is positive in DNA, if it is, triggering third determining module, such as
Fruit is no, then triggers the 4th determining module；
Third determining module, for determining, there are primers in DNA, and primer is known as target primer；
4th determining module, for determining, there is no primers in DNA.
It is described in detail below to the DNA extraction process in cerebrospinal fluid and to the diagnosis process of pathogen.
First, paramagnetic particle method extracts the DNA in Cerebrospinal Fluid in Patients, specifically includes the following steps:
The first step collects 1mL determinand (cerebrospinal fluid) and 1.5mL centrifuge tube 13 is added.Second step is centrifuged at 9000rpm
30 seconds, cell precipitation is made to get off, abandons supernatant, be vortexed or flick and break up cell precipitation.Vortex oscillation until cell mass is sufficiently resuspended,
Dispersion.The resuspension dispersion of cell is extremely important to cracking in next step, and cell, which is not broken up, is just added lysate, and will lead to cell cannot
Sufficiently cracking forms naked eyes visible lumps.Third step is added the cell that 350 μ l Lysis Buffer A are resuspended, blows and beats up and down
Lytic cell, or the 10 seconds help lytic cells that are acutely vortexed, 70 DEG C of water-bath 10min.4th step (in practical operation, executes
Or do not execute the step and do not influenced on result is extracted), RNase A (10mg/ml) is added in lysate to final concentration
10 μ g/ml, overturn 25 mixings, and 37 DEG C of incubation removals in 15 minutes remain RNA, then cool back room temperature.5th step is added ice-cold
150 μ l Solution AB after, on turbula shaker high speed continuous oscillation mix 25 seconds, may be seen after mixing some small
Protein mass.6th step, 12000rpm (can adjust as needed and increase centrifugal force) centrifugation 5 minutes, can see pipe at this time
The lumpy precipitate of bottom crineous or white, it is also possible to see some albumen precipitations and swim in liquid surface.7th step, it is careful to inhale
Take supernatant (about 400 μ l) into a new 1.5ml centrifuge tube 13.When drawing supernatant, be careful not to be drawn onto tube bottom and drift
Float on the albumen precipitation of liquid surface.If be accidentally transferred to albumen precipitation in new centrifuge tube 13, can be centrifuged again 2 minutes
After take supernatant.Isometric 400 μ l of room temperature Binding Buffer PQ and 20 μ l MagDNA magnetic beads is added, at once in 8th step
It turns upside down 20 times and mixes well, at this time it is possible that flocculent deposit, it is also possible to will form one or similar to double helix
Rope cannot siphon away black clumps in practical operation.Centrifuge tube 13 is put into static on magnetic separation rack by the 9th step
1-2min is clarified until solution becomes, and careful sops up liquid, not be drawn onto magnetic bead.Tenth step is removed 1.5ml centrifuge tube 13, is added
Enter 500 μ l impurity removal buffer Wash Buffer B (please first check whether and dehydrated alcohol has been added) to be mixed by inversion, it will
1.5ml centrifuge tube 13 be put into it is 1-2 minutes static on magnetic separation rack, until solution become clarify, carefully sop up liquid.11st
Step is added 600 μ l rinsing liquid Wash Buffer C (please first check whether and dehydrated alcohol has been added), is mixed by inversion, by 1.5ml
Centrifuge tube 13 is put into static 1-2min on magnetic separation rack, clarifies until solution becomes, and careful sops up liquid, discards waste liquid.The
12 steps are added 500 μ lWash Buffer D rinsing liquids (please first check whether and anhydrous second ferment has been added), are mixed by inversion, will be from
Heart pipe 13 is put into static 1-2min on magnetic separation rack, clarifies until solution becomes, and careful sops up liquid, discards waste liquid.(to the greatest extent may be used
Energy sops up residual liquid, in order to avoid influence experimental result).13rd step removes 1.5ml centrifuge tube 13, places it in 37-65
DEG C baking oven in dry residual ethanol.(2-3min can also be blown with hair dryer, should not be too dry, in order to avoid DNA is difficult to dissolve).The
14 steps are added 50-100 μ l elution buffer gently springing tube wall, magnetic bead are broken up completely and is dispersed in elution buffer, room
Temperature places 1-2min, and (elution buffer pre-heat effect in 65-70 DEG C of water-bath is more preferable, if the DNA that obtain higher concentration can
It to reduce elution volume, can also elute in two times).Centrifuge tube 13 is put into static 1-2mim on magnetic separation rack by the 15th step,
It is clarified until solution becomes, careful transfer solution not be drawn onto magnetic bead, if being drawn onto magnetic bead can weigh into a new centrifuge tube 13
New placement separates on magnetic frame.16th step after obtaining DNA extracting solution, can be deposited under 2-8 DEG C of environment, such as
Fruit will store for a long time, can be placed under -20 DEG C of environment.
Second, the DNA purity of extraction is identified.
Because the OD260/OD280 of pure DNA/RNA is 1.8 or 2.0, therefore can be estimated according to the ratio of OD260/OD280
Count out the purity of DNA.If the higher explanation of ratio contains RNA, the lower explanation of ratio is with the presence of residual protein.OD230/OD260
Ratio should be between 0.4-0.5, if the higher explanation of ratio is with the presence of remaining salt.Specifically, if the ratio of OD260/OD280
R is greater than 1.8, illustrates there are RNA, can be handled again with RNaseA.If R value less than 1.8, illustrates have the impurity such as protein to deposit
, need to again with Proteinase K, SDS and expect, chloroform, isoamyl alcohol again purify to DNA (3M of 1/8 volume can also be added
NaAc (pH5.2) promotes DNA Precipitation together with cold ethyl alcohol.
Second, the source of infection in Cerebrospinal Fluid in Patients is detected.
The first step configures primer solution.(4000rpm, 30~60s) is please first centrifuged before uncapping；Slowly open on test tube 14
Helixseal lid 140, suitable buffer solution or water is added；Screw on fully shake after the lid 140 of helixseal it is mixed
It is even；Do not have to temporarily or long term storage needs are placed on -20 DEG C of refrigerations.
Second step, firstly, configuring PCR reaction solution in the way of table 1.
1 PCR reaction solution of table proportion
| Agent formulations | Usage amount | Usage amount | Usage amount |
| qPCR MasterMix | 10ul | 12.5ul | 25ul |
| PCR forward primer | 0.5ul | 0.5ul | 1ul |
| PCR reverse primer | 0.5ul | 0.5ul | 1ul |
| DNA profiling | 1ul | 1ul | 2ul |
| DdH2O (sterile purified water) | 8.0ul | 10.5ul | 21ul |
| It amounts to | 20ul | 25ul | 50ul |
In 20ul reaction system, the additive amount of DNA profiling is usually in 100ng hereinafter, because in different types of DNA profiling
The copy number of the target gene contained is different, can carry out gradient dilution when necessary, determine optimal DNA profiling additive amount.If be intended to
The second step pcr amplification reaction of 2 step Real Time-PCR reaction is carried out using this product, the reaction solution of the first step is as DNA mould
Additive amount when plate does not exceed the 10% of PCR reaction solution total volume.96 orifice plates use 50ul reaction system.
Secondly, carrying out Real Time-PCR reaction.Solubilising reagent, mixing of turning upside down, confirmation reaction solution are completely dissolved
(avoiding mixing using oscillator, acutely oscillation can reduce properties of product)；By configured PCR reaction mixture, packing to PCR
In reaction tube；Add the template of PCR amplification；Pcr amplification reaction is carried out using Real Time PCR amplification instrument；Reaction solution is matched
It sets method and PCR amplification condition please refers to application method；The application method of Real Time PCR instrument, please refers to corresponding instrument
Specification.
In practical applications, the three-step approach PCR response procedures that following display can be used, which diagnose, determines the source of infection, if the journey
Sequence cannot get good experimental result, can carry out the optimization of PCR condition again.Three-step approach PCR amplification standardization program is as follows:
1, initial denaturation: 95 DEG C of 2min.2, PCR reacts, and 10s is carried out under 95 DEG C of environment, 30- is carried out under 56 DEG C of environment
34s carries out 30s under 72 DEG C of environment, and the process is recyclable to be executed 40 times.Annealing temperature can be set according to primer.3, melt
Curve carries out 1min under 95 DEG C of environment, and 1min is carried out under 55 DEG C of environment, and 86 circulations are carried out under 55 DEG C of -98 DEG C of environment.
Finally, Real Time-PCR data are analyzed.The amplification curve of Real Time-PCR is confirmed after reaction and is melted
Solution curve carries out making standard curve etc. when PCR is quantitative.The specific effect using following assessment real-time quantitative PCR reaction,
1, PCR amplification efficiency at least needs to do 3 parallel repetitions, at least does 5 orders of magnitude to correctly assess PCR amplification efficiency
Multiple (5logs) continuous gradient dilutions template concentrations.2, R2 value, it is to illustrate that the statistics of degree of correlation between two values is academic
Language.If R2 is equal to 1, can be with Y value (Ct) come Accurate Prediction X value (amount).If R2 be equal to 0, cannot by Y value come
Predict X value.When R2 value is greater than 0.99, relevant confidence level is fine between two values.3, accuracy, standard deviation be (deviation
Square root) it is most common accuracy metering method.If many data points are all close to average value, standard deviation is with regard to small；
If many data points are all far from average value, standard deviation is with regard to big.In fact, the data group meeting that enough numbers of repetition generate
Form rough normal distribution.This can often prove that I.i.d. random variables exist by classical central limit theory
It is intended to normal distribution when unlimited more.If PCR reaction efficiency is 100%, the average interval Ct between 2 times of dilution points is answered
This is just 1 Ct value.2 times of diluted concentrations are differentiated with 99.7% probability, standard deviation is just necessarily less than equal to 0.167.Mark
Quasi- deviation is bigger, and it is lower to differentiate 2 times of diluted abilities.In order to tell 2 times of dilutions in the case where 95% or more, mark
Quasi- deviation is necessarily less than equal to 0.250.4, sensitivity, no matter CT absolute value is how many, any effectively to expand and detect
The system that beginning template copy numbers are 1 has all reached the limit of sensitivity.PCR efficiency is to determine the key factor of reaction sensitivity.
Another important consideration factor when detecting extremely low copy number is, template number when low-copy cannot be come by general case
It is expected that.On the contrary, it can follow Poisson distribution, that is, carry out a large amount of parallel starting mould that repeats, should averagely copy containing one
Plate, actually about 37% without containing copy, and only about 37% containing 1 copy, and about 18% actually containing there are two copies.Cause
This, in order to more reliably detect low-copy, it is necessary to do a large amount of parallel repetition experiment to provide statistical significance, and overcome Poisson
The limitation of distribution.Evaluate the standard of fluorescent PCR result.
Particularly as be using the first acquisition module 201 according to the first acquisition instruction for receiving obtain using in kit from
The DNA information extracted in the slave cerebrospinal fluid of heart pipe storage recycles second to obtain module according to the second acquisition instruction received
Obtain a variety of primers；Namely by the DNA information extracted and a variety of a variety of of checkout and diagnosis is needed to draw by artificial mode
Object is input to related system, and then, the first determining module 203 can determine target primer present in DNA according to DNA information,
DNA extracting solution and corresponding target primer can be specifically instilled in the reactive tank on 96 hole PCR reaction plates of kit, one anti-
It answers and instills a kind of target primer in slot, the target primer instilled by relevant detecting instrument or method detection is in current reactive tank
Whether it is positive in the reaction solution held, if it is, illustrating that corresponding patient has infected the primer, if the target instilled
Whether primer is negative in the reaction solution that current reactive tank is held, then illustrates that corresponding patient does not infect the primer, weight
This step is executed again, is completed until all target primers are detected, is finally drawn using the second determining module 204 according to target
Object determines corresponding antibiotic to diagnose to the pathogen in cerebrospinal fluid；Wherein, primer includes target primer.Work as determination
Out after the source of infection of current patents, so that it may accurately carry out medication to the patient, improve the therapeutic efficiency of patient.
A kind of cerebrospinal fluid difficulty pathogen diagnostic device provided herein, the first acquisition module can be according to receiving
The first acquisition instruction obtain and utilize the DNA information extracted in kit in the slave cerebrospinal fluid of centrifuge tube storage；Second obtains mould
Block can obtain a variety of primers according to the second acquisition instruction received；First determining module can determine DNA according to DNA information
Present in target primer；Second determining module can determine corresponding antibiotic in cerebrospinal fluid according to target primer
Pathogen is diagnosed；Wherein, primer includes target primer.That is, the diagnostic device may be implemented to a variety of primers
Diagnosis detection, i.e. actual needs detection how many kinds of primer (source of infection), so that it may realize to the checkout and diagnosis in multi-infection source, mention
The detection accuracy of the high source of infection, and then may insure clinical precisely diagnosis pathogen infection type and adjustment pesticide application strategy, drop
Low mortality and shortening patient caused by infection because being hospitalized the period.
On the basis of the above embodiments, preferably embodiment, the type of primer are specially 20 kinds, respectively greatly
The uncommon bacterium of intestines angstrom, enterococcus faecalis, streptococcus pneumonia, serratia marcescens, proteus mirabilis, stenotrophomonas maltophilia, Bao Man
Acinetobacter calcoaceticus, pseudomonas aeruginosa, enterococcus faecium, klebsiella pneumoniae, staphylococcus aureus, Bu Shi citric acid bacillus,
Burkholderia cepacia, staphylococcus epidermis, Candida albicans, Candida tropicalis, Candida parapsilosis bacterium, smooth false silk
Saccharomycete, candida krusei, aspergillus fumigatus, herpe simplex, HHV-6, CMV and VZV.
Have chosen 25 kinds of primers (source of infection or bacterium) in the embodiment of the present application, be specifically shown in Table 2,25 kinds of primers it is detailed
Situation is as shown in table 2.Wherein, including 14 kinds of bacteriums, 7 kinds of fungies, 4 kinds of viruses.
The details of each bacterium of table 2
Those skilled in the art will readily occur to its of the application after considering specification and practicing application disclosed herein
His embodiment.This application is intended to cover any variations, uses, or adaptations of the application, these modifications, purposes or
Person's adaptive change follow the general principle of the application and include common knowledge in the art disclosed in the present application or
Conventional techniques.Description and embodiments are considered only as exemplary, and the just true range of the application is pointed out by claim.
It should be understood that the application is not limited to the precise structure that has been described above and shown in the drawings, and
And it can carry out various modifications and change without departing from the scope again.Above-described the application embodiment is not constituted to this Shen
Please protection scope restriction.
Claims (10)
1. a kind of kit for cerebrospinal fluid difficulty pathogen diagnosis, including reagent box body, which is characterized in that further include:
Positioned at the fixed plate of the reagent box body, primer plate and 96 hole PCR reaction plates；
The first groove for fixing the primer plate and second for placing each centrifuge tube recessed is provided in the fixed plate
Slot, the number of second groove and the number of the centrifuge tube correspond；
The third groove for placing multiple test tubes that plurality of target primer is housed is provided on the primer plate, the third is recessed
The number of slot and the number of the test tube correspond, and a test tube is equipped with a kind of target primer, and the test tube goes out
The lid of helixseal is installed at mouthful.
2. the kit according to claim 1 for cerebrospinal fluid difficulty pathogen diagnosis, which is characterized in that the spiral
The sealing ring for sealing is additionally provided on the lid of sealing.
3. the kit according to claim 1 for cerebrospinal fluid difficulty pathogen diagnosis, which is characterized in that each described
Two grooves are located at the same side of the fixed plate.
4. the kit according to claim 3 for cerebrospinal fluid difficulty pathogen diagnosis, which is characterized in that each described
Two grooves are uniformly distributed in the same side of the fixed plate.
5. the kit according to claim 1 for cerebrospinal fluid difficulty pathogen diagnosis, which is characterized in that the primer
The cross section of plate is rectangle；
Accordingly, the cross section of first groove is rectangle.
6. the kit according to claim 5 for cerebrospinal fluid difficulty pathogen diagnosis, which is characterized in that further include:
It is installed on the primer plate, the lid being engaged positioned at the edge of third groove side and the primer plate.
7. a kind of cerebrospinal fluid difficulty pathogen diagnostic device, based on doubtful for cerebrospinal fluid described in claim 1 to 6 any one
The kit of difficult pathogen diagnosis characterized by comprising
First obtains module, for obtaining the slave brain using centrifuge tube storage in kit according to the first acquisition instruction received
The DNA information extracted in spinal fluid；
Second obtains module, for obtaining a variety of primers according to the second acquisition instruction received；
First determining module, for determining target primer present in DNA according to the DNA information；
Second determining module, for determining corresponding antibiotic to the cause of disease in the cerebrospinal fluid according to the target primer
Body carries out diagnosis and recommends positive case to apply corresponding antibiotic treatment；
Wherein, the primer includes the target primer.
8. cerebrospinal fluid difficulty pathogen diagnostic device according to claim 7, which is characterized in that first determining module
Specifically for determining the target primer by PCR-Array mode.
9. cerebrospinal fluid difficulty pathogen diagnostic device according to claim 8, which is characterized in that first determining module
It specifically includes:
Judgment module, for judging whether each primer is positive in the DNA, if it is, triggering third determines mould
Block, if it is not, then the 4th determining module of triggering；
Third determining module, for determining, there are the primers in the DNA, and the primer is known as the target primer；
4th determining module, for determining, there is no the primers in the DNA.
10. according to cerebrospinal fluid difficulty pathogen diagnostic device described in claim 7 to 9 any one, which is characterized in that described
The type of primer is specially 25 kinds, respectively escherichia coli, enterococcus faecalis, streptococcus pneumonia, serratia marcescens, unusual change
Shape bacillus, stenotrophomonas maltophilia, Acinetobacter bauamnnii, pseudomonas aeruginosa, enterococcus faecium, klebsiella pneumoniae, gold
Staphylococcus aureus, Bu Shi citric acid bacillus, Burkholderia cepacia, staphylococcus epidermis, Candida albicans, tropical beads
Bacterium, Candida parapsilosis bacterium, Candida glabrata, candida krusei, aspergillus fumigatus, herpe simplex, HHV-6, CMV and
VZV。
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| CN112249488A (en) * | 2020-10-09 | 2021-01-22 | 深圳市森盈生物科技有限公司 | Kit for diagnosing cerebrospinal fluid difficult pathogen and diagnostic device thereof |
Citations (14)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20020095073A1 (en) * | 2000-11-27 | 2002-07-18 | Jacobs Alice A. | Clinically intelligent diagnostic devices and mehtods |
| US20040010129A1 (en) * | 2001-11-01 | 2004-01-15 | Po-Hsiang Hsu | Nucleic acid kit for bacterial pathogen diagnosis and method for using the same |
| CN101048513A (en) * | 2004-08-25 | 2007-10-03 | 阿里法克斯技术有限公司 | Integrated device for diagnostic analyses, and relative method |
| CN101144775A (en) * | 2007-08-31 | 2008-03-19 | 浙江大学 | Bacteria real-time fluorescence quantitative polymerase chain reaction detection reagent kit and its uses |
| CN102150032A (en) * | 2008-09-04 | 2011-08-10 | 索尼Dadc公司 | Diagnostic system with portable collection housing |
| US20120100546A1 (en) * | 2010-10-22 | 2012-04-26 | T2 Biosystems, Inc. | Nmr systems and methods for the rapid detection of analytes |
| CN102770559A (en) * | 2009-12-03 | 2012-11-07 | 奎斯特诊断投资有限公司 | Methods for the diagnosis of bacterial vaginosis |
| US20130157301A1 (en) * | 2010-07-26 | 2013-06-20 | Assistance Publique - Hopitaux De Paris | Diagnosis of bacterial meningitis based on the measure of ROS production in a sample of cerebrospinal fluid |
| CN105143435A (en) * | 2013-02-28 | 2015-12-09 | 远程医疗零距离公司 | Diagnostic device for evaluating microbial content of a sample |
| CN206273830U (en) * | 2016-08-08 | 2017-06-23 | 深圳市检验检疫科学研究院 | Human corona virus's NL63 detection kits |
| CN207109001U (en) * | 2017-08-29 | 2018-03-16 | 王碧 | A kind of novel agent box |
| CN108431237A (en) * | 2015-09-10 | 2018-08-21 | 弗劳恩霍夫应用研究促进协会 | For the unusual condition including determining sample, the especially method and apparatus of the diagnostic method based on nucleic acid of health status and/or pathogenic situation |
| CN108728543A (en) * | 2018-06-22 | 2018-11-02 | 南阳师范学院 | Detect the miRNA combination of brain metastasis and the kit containing the combination |
| CN209636254U (en) * | 2019-01-09 | 2019-11-15 | 赵志军 | A kind of kit for cerebrospinal fluid difficulty pathogen diagnosis |
-
2019
- 2019-01-09 CN CN201910018294.3A patent/CN109439737A/en active Pending
Patent Citations (15)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20020095073A1 (en) * | 2000-11-27 | 2002-07-18 | Jacobs Alice A. | Clinically intelligent diagnostic devices and mehtods |
| US20040010129A1 (en) * | 2001-11-01 | 2004-01-15 | Po-Hsiang Hsu | Nucleic acid kit for bacterial pathogen diagnosis and method for using the same |
| CN101048513A (en) * | 2004-08-25 | 2007-10-03 | 阿里法克斯技术有限公司 | Integrated device for diagnostic analyses, and relative method |
| CN101144775A (en) * | 2007-08-31 | 2008-03-19 | 浙江大学 | Bacteria real-time fluorescence quantitative polymerase chain reaction detection reagent kit and its uses |
| CN102150032A (en) * | 2008-09-04 | 2011-08-10 | 索尼Dadc公司 | Diagnostic system with portable collection housing |
| CN102770559A (en) * | 2009-12-03 | 2012-11-07 | 奎斯特诊断投资有限公司 | Methods for the diagnosis of bacterial vaginosis |
| US20130157301A1 (en) * | 2010-07-26 | 2013-06-20 | Assistance Publique - Hopitaux De Paris | Diagnosis of bacterial meningitis based on the measure of ROS production in a sample of cerebrospinal fluid |
| US20120100546A1 (en) * | 2010-10-22 | 2012-04-26 | T2 Biosystems, Inc. | Nmr systems and methods for the rapid detection of analytes |
| CN105143435A (en) * | 2013-02-28 | 2015-12-09 | 远程医疗零距离公司 | Diagnostic device for evaluating microbial content of a sample |
| CN108431237A (en) * | 2015-09-10 | 2018-08-21 | 弗劳恩霍夫应用研究促进协会 | For the unusual condition including determining sample, the especially method and apparatus of the diagnostic method based on nucleic acid of health status and/or pathogenic situation |
| US20180307795A1 (en) * | 2015-09-10 | 2018-10-25 | Fraunhofer-Gesellschaft zur Förderung der angewandten Forschung e.V. | Method and device for nucleic acid based diagnostic approaches including the determination of a deviant condition, especially a health condition and/or a pathogenic condition of a sample |
| CN206273830U (en) * | 2016-08-08 | 2017-06-23 | 深圳市检验检疫科学研究院 | Human corona virus's NL63 detection kits |
| CN207109001U (en) * | 2017-08-29 | 2018-03-16 | 王碧 | A kind of novel agent box |
| CN108728543A (en) * | 2018-06-22 | 2018-11-02 | 南阳师范学院 | Detect the miRNA combination of brain metastasis and the kit containing the combination |
| CN209636254U (en) * | 2019-01-09 | 2019-11-15 | 赵志军 | A kind of kit for cerebrospinal fluid difficulty pathogen diagnosis |
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| CN112249488A (en) * | 2020-10-09 | 2021-01-22 | 深圳市森盈生物科技有限公司 | Kit for diagnosing cerebrospinal fluid difficult pathogen and diagnostic device thereof |
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