CN107589269A - Detection card for A type antibodies against foot-and-mouth disease virus in Quantitative detection serum - Google Patents
Detection card for A type antibodies against foot-and-mouth disease virus in Quantitative detection serum Download PDFInfo
- Publication number
- CN107589269A CN107589269A CN201711054389.8A CN201711054389A CN107589269A CN 107589269 A CN107589269 A CN 107589269A CN 201711054389 A CN201711054389 A CN 201711054389A CN 107589269 A CN107589269 A CN 107589269A
- Authority
- CN
- China
- Prior art keywords
- mouth disease
- detection
- disease virus
- foot
- type
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
Landscapes
- Investigating Or Analysing Biological Materials (AREA)
Abstract
The invention discloses a kind of detection card for being used for A type antibodies against foot-and-mouth disease virus in Quantitative detection serum, including detection card shell and the test strips being assemblied in detection card shell, the test strips include the plastic bottom board with pressure sensitive adhesive, paste sample pad successively on bottom plate, labeling pad, nitrocellulose filter and blotting paper, the labeling pad is made up of carrier substrate and label, label is to spray lanthanide fluoro in carrier substrate to detect the tunic that microballoon and lanthanide fluoro Quality Control microballoon are formed, it is detection line that A type foot and mouth disease virus epitope recombinant antigens are coated with the nitrocellulose filter, it is nature controlling line to be coated with rabbit-anti chicken lgY antibody, label is the fluorescence Quality Control microballoon of the fluoroscopic examination microballoon and mark chicken lgY antibody that are marked with A type FMDV VP1 recombinant antigens.The achievable A type antibodies against foot-and-mouth disease virus scene Quantitative detection of the present invention, real-time monitoring and evaluation immune effect of vaccine.
Description
Technical field
The present invention relates to one kind detection card and use, be used for A type mouths in Quantitative detection serum more particularly to one kind
The detection card of aphtovirus antibody.
Background technology
Aftosa（Foot-and-Mouth Disease, FMD）It is by foot and mouth disease virus（Foot-and-Mouth
Disease Virus, FMDV）Caused pig, ox, the great animal epidemic of sheep main economic poultry kind morbidity.The sick route of transmission
It is more, speed is fast, epidemic regions are wide, host's spectrum width, once morbidity not only causes huge economic loss, and can cause severe
Political fallout, enjoy countries in the world to pay high attention to, be known as the title of " political economy disease "., OIE（Office
International des Epizooties, OIE) animal epidemic that must circulate a notice of is classified as, China is classified as A classes and moved
First of thing infectious disease.With the quickening of the global trade process of integration, trans-regional, a wide range of quick biography is also presented in aftosa prevalence
The situation broadcast, prevention and control become increasingly difficult.China's aftosa prevention and control situation also allows of no optimist, and mainly China boundary line is very long,
With many countries that China borders on because aftosa epidemic situation occurs for the factors such as politics, economic, culture and war throughout the year, especially print
The degree peninsula is known as the title in " foot and mouth disease virus storehouse ", and China's aftosa prevention and control are caused with very big pressure.Most typically 2009
Year is 2 different topological types with incoming A types aftosa in 2013, and huge economic damage is caused to China's livestock breeding industry
Lose.
At present, aftosa prevention and control in China's mainly use vaccine immunity, the prevention and control being combined with slaughtering infection and suspicious drove
Strategy.Animal population protection antibody level for prevention infection of foot-and-mouth disease serve it is conclusive, therefore, real-time monitor
Animal population immune antiboidy is horizontal, formulates scientific and effective immune programme for children according to Monitoring Data, strengthens animal population immune antiboidy
It is horizontal most important to effective prevention and control aftosa.
Nowadays A types foot-and-mouth disease antibody is detected still based on qualitative/quantitative ELISA kit, is China's aftosa prevention and control
Provide important technical support, but the technology still has the shortcomings that many and deficiency, first, complex operation, labor intensity is big；
Secondly, it is necessary to specialized laboratories and professional's operation；Furthermore experimental period is grown, and it is slow to go out result.Foot-and-mouth disease antibody detects colloid
Gold test paper strip is although easy to detect, is not required to professional and technical personnel and working environment；But its sensitivity detected is low, and can only determine
Property, detection range is narrow.Therefore, development is quick, convenient, can carry out the fast of quantitative detection antibodies against foot-and-mouth disease virus with field at the scene
Fast detection technique is particularly important.
The content of the invention
It is used to A types antibodies against foot-and-mouth disease virus in Quantitative detection serum it is an object of the invention to provide one kind detect
Card, overcomes collaurum rapid detection card and Kit each shortcoming, by shirtsleeve operation, can at the scene with field pair
Pig, ox, sheep and other animal blood serum samples carry out it is quick, accurate, quantitatively detect with sensitivity, the technology is both with ELISA
Sensitiveness, there is the convenience of colloidal gold strip again, it may be said that fundamentally solve above-mentioned the deficiencies in the prior art part.
The purpose of the present invention is realized by following technical proposals：
A kind of detection card for being used for A type antibodies against foot-and-mouth disease virus in Quantitative detection serum, including detection card shell and assembling
Test strips in detection card shell, the test strips include the plastic bottom board with pressure sensitive adhesive, paste sample successively on bottom plate
Pad, labeling pad, nitrocellulose filter and blotting paper, the labeling pad are made up of carrier substrate and label, and label is
The tunic that lanthanide fluoro detection microballoon and lanthanide fluoro Quality Control microballoon are formed, the nitrocellulose filter are sprayed in carrier substrate
Upper coating A type foot and mouth disease virus epitope recombinant antigens are detection line, and coating rabbit-anti chicken lgY antibody is nature controlling line, and label is mark
Note has the fluoroscopic examination microballoon of A type FMDV VP1 recombinant antigens and the fluorescence Quality Control microballoon of mark chicken lgY antibody.
Further, the foot and mouth disease virus epitope recombinant antigen is the epitope antigen of A type FMDV VP1s antigenic region,
It is the epitope recombinant protein of expression.
Further, the rabbit-anti chicken lgY antibody that nature controlling line uses on the nitrocellulose filter for rabbit-anti chicken lgY lgG.
A kind of method for preparing above-mentioned A types Foot-and-mouth disease, using bioinformatics technique to A type aftosas
The VP1 antigen genes of the different lineage strains of virus are compared, design synthesis A type FMDV VP1 antigen genes, and
Prokaryotic expression plasmid structure A type foot-and-mouth disease virus antigen DNA recombinant expression plasmids are cloned into, are then converted large intestine
Bacillus competent cell BL21（DE3）PLysS screening recombination expression bacterium, recombination expression bacterium is through IPTG induced expression A type aftosas
Virus VP 1 recombinant antigen, obtain detection through affinitive layer purification and use VP1 recombinant antigens.
A kind of method for preparing above-mentioned A types foot and mouth disease virus epitope antigen albumen, should using bioinformatics technique principle
A type FMDV VP1 antigen genes are compared with computer, predict epitope, screening and design A type mouth hoof
Epidemic disease poison epitope antigen gene, and it is cloned into prokaryotic expression plasmid structure A type foot and mouth disease virus epitope antigen genetic recombination
Expression plasmid, then converted competent escherichia coli cell BL21（DE3）PLysS screening recombination expression bacterium, recombination expression
Bacterium obtains another detection antigen through IPTG induced expression A type foot and mouth disease virus epitope recombinant antigens through affinitive layer purification.
A kind of application method of above-mentioned detection card, the detection card are quantitative chromatography detection card, will detect blood sample prepare liquid
Instill in the well of detection card, chromatograph 15 minutes, then detection card is scanned using chromatogram scanner, chromatogram scanner
Scanning the data obtained is radioed into client, the upper nature controlling line of detection card and detection is calculated according to scan data in client
The fluorescence signal intensity value of line, while client obtains detected A types antibodies against foot-and-mouth disease virus standard curve, client from cloud platform
A types mouth in prepare liquid is calculated according to the contrast relationship of standard curve and nature controlling line and the fluorescence signal intensity value of detection line in end
The antibody titer of aphtovirus antibody, judge test sample result.
Preferably, the standard curve is to import cloud platform in the following way, a collection of detection card is prepared first, and
A types antibodies against foot-and-mouth disease virus standard items corresponding to preparation, block chromatography standard items with detection and detected with chromatogram scanner, chromatography is swept
Retouch instrument and the data processing that detection obtains is passed into client, detection line corresponding to standard items and nature controlling line is calculated in client
Fluorescence signal intensity value, and returned according to this data so as to obtain A type antibodies against foot-and-mouth disease virus standard curves, client
Standard curve is transmitted to cloud platform and stored.
Preferably, the standard curve that the client is calculated forms a file, and corresponding generation bar code, visitor
Family end can extract the standard curve by scanning bar code from cloud platform.
Explanation of nouns：
Rabbit-anti chicken lgY lgG：Rabbit-anti chicken lgY immunoglobulin G
Chicken lgY antibody：Chicken yolk antibody
Competent escherichia coli cell BL21（DE3）：The most frequently used host strain of escherichia expression system,
IPTG：Isopropyl-β-D-thiogalactoside.
Compared with prior art, the beneficial effects of the present invention are：High sensitivity, batch internal difference and difference between batch of detection are small,
There is preferable repeatability, stable performance, both can detect whole blood sample, serum and plasma sample can also be detected, with CSFV（Swine fever
Virus）、PRRSV（Pig breathe with breeding dysfunction syndrome otopathy virus), PCV（Pig circular ring virus）、PRV（Pseudorabies virus),
PPV（Pig parvoviral）、JEV（Encephalitis B virus) etc. antibody there is no cross reaction.It is fast that A types antibodies against foot-and-mouth disease virus scene can be achieved
Speed detection, will provide high-quality and efficient, fast and convenient monitoring means and technology, significant increase China mouth for China's aftosa prevention and control
Fever aphthous Control Technology is horizontal, has higher practical value and promotional value, and the recombinant protein expressed using multi-epitope can be to prevent
Leak-stopping is examined, and improves detection in susceptibility.
Brief description of the drawings
Fig. 1 is the structural representation of the present invention；
Fig. 2 is the structural representation of test strips in Fig. 1；
Fig. 3 is Fig. 2 top view；
Fig. 4 is A type antibodies against foot-and-mouth disease virus examination criteria curves；
Fig. 5 is another structural representation of present invention detection card.
Embodiment
With reference to specific embodiments and the drawings, the present invention is further illustrated.
As shown in Figures 1 to 4, a kind of detection card for being used for A type antibodies against foot-and-mouth disease virus in Quantitative detection serum, bag
The test strips being assemblied in shell 10 are included, the test strips include the plastic bottom board 1 with pressure sensitive adhesive, from a left side on plastic bottom board 1
Sample pad 2, labeling pad 3, nitrocellulose filter 4 and blotting paper 5 are pasted successively to right, and the right-hand member of sample pad 2 rides over mark
On the left end of thing pad 3, the right-hand member of labeling pad 3 is ridden on the left end of nitrocellulose filter 4, and the left end of blotting paper 5 rides over nitric acid fibre
On the right-hand member for tieing up plain film 4.It is detection line 6 that A type foot and mouth disease virus epitopes recombinant antigen is coated with the nitrocellulose filter 4, bag
It is nature controlling line 7 by rabbit-anti chicken lgY antibody, counter sample pad 2 opens up well 11, corresponding nitrocellulose filter 4 on shell 10
On detection line 6 and nature controlling line 7 open up form hole 12.
The A types foot and mouth disease virus epitope recombinant antigen is the Main Antigenic of A type FMDV VP1s antigenic region,
It is the epitope recombinant protein of expression.
The rabbit-anti chicken lgY antibody that nature controlling line uses on the nitrocellulose filter 4 for rabbit-anti chicken lgY lgG.
The labeling pad 3 is made up of carrier substrate and label, and label is to be marked with A type FMDV VP1 weights
The fluoroscopic examination microballoon of group antigen and the fluorescence Quality Control microballoon of mark chicken lgY antibody.A types antibodies against foot-and-mouth disease virus, A type aftosas
It is by double that A type foot and mouth disease virus epitope recombinant antigens are coated with the label and nitrocellulose filter of virus VP 1 recombinant antigen
A type antibodies against foot-and-mouth disease virus in antigen sandwich method detection pig, ox, sheep and other animal blood samples, judge to tie according to antibody titer
Fruit.
As a kind of optimal technical scheme, it is described detection card the surface of shell 10 correspond to blotting paper 5 position offer it is more
Individual through hole 13, be advantageous to solvent and volatilize shell 10, it is ensured that testing result is accurate, and the detection card shell for solving prior art is adopted
The technical problem that can not be volatilized away with enclosed construction, solvent, referring to Fig. 5.
In order to ensure volatilization effect, equidistantly uniformly opened in the region that the surface of shell 10 matches with the size of blotting paper 5
If through hole 13.
As another optimal technical scheme, the detection card body, which tilts, to be fixed in shell 10, and sample pad 2 is in
Low side, blotting paper 5 are in high-end.This structure causes the testing sample reduced velocity flow instilled from well 11, slowly toward upper strata
Analysis, ensures the enough chromatography time, can effectively prevent that liquid sample flow is too fast and influences testing result, avoid the too fast stream of sample
Entering nitrocellulose filter 4 causes testing result to fail.Because if testing sample flow velocity is too fast, easily cause to chromatograph it is insufficient,
And then influence testing result.The detection inclined angle of card body is specifically designed according to the difference of testing sample.
As another preferred structure of chromatography detection card, holding tank is provided with the shell 10 of the chromatography detection card, and
2 holding tanks are arranged with the both sides of detection card body, sheet drier is installed in holding tank.This design causes dry
Drying prescription keeps together with detection card body so that detection card body humidity is effectively controlled, so as to improve accuracy of detection.
As another preferred structure of chromatography detection card, can also be set on the surface of shell 10 of chromatography detection card with cover
Sample accumulator tank, sample can be sealed up for safekeeping.The hd top face of the sample accumulator tank flushes with the surface of shell 10, covers provided with recessed
Hole, convenient use person's finger force is turned-out to uncap.
The prokaryotic expression of embodiment one, A type Foot-and-mouth disease VP1 albumen
A type foot and mouth disease virus difference lineage strain VP1 antigen genes are compared using bioinformatics technique, designed
A type foot and mouth disease virus specificity VP1 antigen genes are synthesized, in order to ensure that VP1 antigen genes are correctly inserted into expression vector, in gene
Both ends introduce I/xho of specific cleavage site BamH I, the gene entrusts precious bioengineering（Dalian）Co., Ltd synthesizes,
And pUC-19T carriers are cloned into, it is named as pUC-VP1.With the enzymes of I/xho of BamH I respectively to pUC-VP1 and recombination expression matter
Grain pET-28a（+）Digestion is carried out, with glue purification QIAquick Gel Extraction Kit（Takara, Dalian）To VP1 genetic fragments and pET-28a
（+）DNA fragmentation is reclaimed, then with two fragment connection construction recombination plasmid pET-VP1 of T4 ligases.Turned with connection product
Change BL21（DE3）PLysS competent cells, through containing kanamycins（Kan+）LAB screening positive restructuring bacterium.Select positive weight
Group clone bacterium adds 100ml and contained in Kan+ LB nutrient solutions, 37 DEG C of 220rpm shaken cultivations, works as OD600During=0.4-0.6, add
After the IPTG for entering final concentration of 0.4Mm, continue to cultivate 4-6 hours, culture is collected by centrifugation, after precipitation is resuspended with appropriate PBS, warp
Ultrasonication is handled, and 10000g centrifugation 30min, supernatant is precipitated and carries out SDS-PAGE electrophoresis（Polyacrylamide gel electricity
Swimming）, as a result show, recombinant protein is expressed with soluble form, and expression antigen, its purity are purified by Ni-NTA affinity columns
Up to more than 95%.Western blot（Western blot test）As a result show, recombinant protein can be with A type foot and mouth disease virus positive bloods
It is clear that specific immune response occurs.Detection antigen is used as using this recombinant antigen.
Embodiment two, the expression of A type foot and mouth disease viruses epitope antigen
A kind of method for preparing above-mentioned A types foot and mouth disease virus epitope antigen, using bioinformatics technique to A type foot and mouth disease viruses
Different lineage strain VP1 antigen genes are compared, the Primary epitope antigenic site of design synthesis A type FMDV VP1s
Cause, in order to ensure that epitope antigen gene is correctly inserted into expression vector, specific cleavage site EcoR is introduced at the both ends of gene
I/xho I, the gene entrust precious bioengineering（Dalian）Co., Ltd synthesizes, and is named as AE.Distinguish pairing with the enzymes of I/xho of EcoR I
Into Gene A E and recombinant expression plasmid pGEM-6P-1 carry out digestion, with DNA fragmentation QIAquick Gel Extraction Kit（Takara, Dalian）
Genetic fragment and pGEM-6P-1 DNA fragmentation are reclaimed, then with two fragment connection construction recombination plasmids of T4 ligases
pGEM-AE.Connection product is converted into BL21（DE3）PLysS competent cells, through containing ampicillin（Amp+）LAB sieve
Select positive restructuring bacterium.Select Positive recombinant clones bacterium to add in the LB nutrient solutions that 100ml contains Amp+, 37 DEG C of 220rpm vibrations
Culture, works as OD600During=0.4-0.6, after adding final concentration of 0.6Mm IPTG, continue to cultivate 4-6 hours, culture is collected by centrifugation
Thing, after precipitation is resuspended with appropriate PBS, handled through ultrasonication, 10000g centrifugation 30min, supernatant is precipitated and carries out SDS-
PAGE electrophoresis（Polyacrylamide gel electrophoresis）, as a result show, recombinant protein is expressed with soluble form, passes through affinity column
Purifying expression antigen, its purity is up to more than 95%.Western blot（Western blot test）As a result show, recombinant protein energy and A
Specific immune response occurs for type foot and mouth disease virus positive serum.Alternatively detected with this epitope recombinant antigen with anti-
It is former.
Embodiment three, prepare A types antibodies against foot-and-mouth disease virus detection card
Prepare A type antibodies against foot-and-mouth disease virus examination criteria curves：Configure the calibration solution containing A type antibodies against foot-and-mouth disease virus（Type containing A
Antibodies against foot-and-mouth disease virus standard items）6 parts, concentration is respectively 0,1/4,1/16,1/64,1/256,1/1024（A type foot and mouth disease viruses
Antibody standard substance doubling dilution）.The calibration solution of above-mentioned various concentrations is separately added into the well of the detection card assembled, layer
After analysis 15 minutes, detected by chromatogram scanner, the testing result that 6 times are obtained is calculated by client process, client
Go out detection line corresponding to standard items and the fluorescence signal intensity value of nature controlling line, and carry out linear regression according to this data and make A types
Antibodies against foot-and-mouth disease virus standard curve, the standard curve that client is calculated form a file, and corresponding generation bar code,
Client is by the file using bar code as filename（Include standard curve）Transmit to cloud platform and store, referring to Fig. 4.
The standard curve that the client is calculated forms a file, and corresponding generation bar code, client pass through
Scanning bar code can extract the standard curve from cloud platform.
The application method of the detection card：By measuring samples（By taking serum as an example）1 is pressed with sample diluting liquid:50 dilution proportions,
The sample 80ul diluted is added in well 11, in the presence of blotting paper 5, sample is from sample pad 2 to the direction of blotting paper 5
It is mobile.15min is chromatographed in the case where avoiding strong illumination, then detection card is detected with chromatogram scanner, chromatography is swept
The fluorescence signal that instrument obtains detection line 6 and nature controlling line 7 is retouched, detection data are passed to client by chromatogram scanner by bluetooth, visitor
Family end calculates the fluorescence signal intensity value of detection line and the fluorescence signal intensity value of nature controlling line according to detection data, and client is led to
The bar code for over-scanning this batch of detection card obtains the standard curve of detected A types antibodies against foot-and-mouth disease virus, client from cloud platform
A types mouth hoof in prepare liquid is calculated according to the contrast relationship of standard curve and nature controlling line and the fluorescence signal intensity value of detection line
Epidemic disease virus antibody titer, referring to Fig. 4 standard curve.Client can be mobile phone or tablet personal computer.
The preparation method of above-mentioned detection card is as follows：
Step 1, nitrocellulose filter is pasted onto on PVC bottom plates, using a film metal spraying special purpose machinery on nitrocellulose filter
The lgG that spraying has been diluted to 0.25mg/ml goat-anti chicken lgY forms nature controlling line and has been diluted to 0.5mg/ml A type aftosas
Virus epitopes recombinant protein forms detection line, and discharge rate is 1 μ l/cm, is then baked 8 hours at a temperature of 37 DEG C；
Step 2, preparation are marked with chicken lgY lanthanide fluoro microballoon and are marked with the lanthanum of A type FMDV VP1 recombinant antigens
It is fluorescent microsphere, 1mL lanthanide fluoro microballoon is added to 50mg MES（2-（N- morpholines）Ethyl sulfonic acid）Buffer solution（0.1M,
pH7.0）In, add 10mg carbodiimides（EDC）With 10mg n-hydroxysuccinimide sulfonate sodium stirring and dissolvings, room temperature
Reaction carries out centrifugally operated after 30 minutes, by centrifugal sediment 50mM borate buffers（pH8.2）Redissolve, add 2mg dialysis
The chicken lgY crossed, stirring reaction 24 hours at ambient temperature, after being then centrifuged for, closing, then preserved in dilution（Preserve ring
Border temperature is 2~8 DEG C）, produce the lanthanide fluoro microballoon for being marked with chicken lgY；A type aftosas are marked using above-mentioned same method
The lanthanide fluoro microballoon of virus VP 1 recombinant antigen；
Step 3, the lanthanide fluoro microballoon and difference for being marked with chicken lgY He being marked with A type FMDV VP1 recombinant antigens
The μ g/ml of concentration 0.1 and 3 μ g/ml are diluted to, is sprayed on using a film metal spraying machine in carrier substrate and forms label pad, is sprayed
Measure as 2.5 μ l/cm, then baked 8 hours at a temperature of 37 DEG C；
Step 4, sample pad, label pad, blotting paper are bonded on PVC bottom plates successively, are assembled into kilocalorie, then cut with cutter
The test strips wide into 5mm, it is assemblied in detection card shell, that is, obtains this A types antibodies against foot-and-mouth disease virus detection card.
The foregoing is merely illustrative of the preferred embodiments of the present invention, is not intended to limit the invention, all essences in the present invention
All any modification, equivalent and improvement made within refreshing and principle etc., should be included in the scope of the protection.
Claims (8)
1. a kind of detection card for being used for A type antibodies against foot-and-mouth disease virus in Quantitative detection serum, including detection card shell and dress
The test strips in detection card shell are fitted over, the test strips include the plastic bottom board with pressure sensitive adhesive, paste sample successively on bottom plate
Product pad, labeling pad, nitrocellulose filter and blotting paper, the labeling pad are made up of carrier substrate and label, label
The tunic that microballoon and lanthanide fluoro Quality Control microballoon are formed is detected to spray lanthanide fluoro in carrier substrate, it is characterised in that：Institute
It is detection line to state and A type foot and mouth disease virus epitope recombinant antigens are coated with nitrocellulose filter, and coating rabbit-anti chicken lgY antibody is Quality Control
Line, label are the fluorescence of the fluoroscopic examination microballoon and mark chicken lgY antibody that are marked with A type FMDV VP1 recombinant antigens
Quality Control microballoon.
2. the detection card according to claim 1 for being used for A type antibodies against foot-and-mouth disease virus in Quantitative detection serum, it is special
Sign is：The A types foot and mouth disease virus epitope recombinant antigen is A type FMDV VP1s area epitope antigen, and it is the table of expression
Position recombinant antigen protein.
3. the detection card according to claim 1 for being used for A type antibodies against foot-and-mouth disease virus in Quantitative detection serum, it is special
Sign is：The rabbit-anti chicken lgY antibody that nature controlling line uses on the nitrocellulose filter for rabbit-anti chicken lgY lgG.
A kind of 4. method for preparing the FMDV VP1 epitope recombinant antigens of A types described in claim 2, it is characterised in that：Adopt
A type foot and mouth disease virus difference lineage strain VP1 antigen genes are compared with bioinformatics technique, design synthesis A
Type foot and mouth disease virus specificity VP1 antigen genes, and be cloned into prokaryotic expression plasmid structure A type FMDV VP1s and resisted
Protogene recombinant expression plasmid, then converted competent escherichia coli cell BL21（DE3）PLysS screening recombination expressions
Bacterium, recombination expression bacterium are after affinitive layer purification through IPTG induced expression A type FMDV VP1 recombinant antigens, antigen
Detect antigen.
A kind of 5. method for preparing the FMDV VP1 epitope recombinant antigens of A types described in claim 2, it is characterised in that：Adopt
With bioinformatics technique principle, the epitope antigen gene of A type foot and mouth disease viruses is compared appliance computer, design
A type foot and mouth disease virus epitope antigen genes are synthesized, and is cloned into prokaryotic expression plasmid structure A type foot and mouth disease virus epitopes and resisted
Protogene recombinant expression plasmid, then converted competent escherichia coli cell BL21（DE3）PLysS screening recombination expressions
Bacterium, recombination expression bacterium are after affinitive layer purification through IPTG induced expression A type foot and mouth disease virus epitope recombinant antigens, antigen
Another detection antigen.
A kind of 6. application method of the detection card of claim 1 or 2 or 3 or 4, it is characterised in that：The detection card is quantitative
Chromatography detection card, it will instill in the well of detection card, chromatograph 15 minutes, then using computed tomography scanning containing test serum sample
Instrument is scanned to detection card, and chromatogram scanner will scan the data obtained and radio to client, and client is according to scanning number
According to the fluorescence signal intensity value that the upper nature controlling line of detection card and detection line is calculated, at the same client obtained from cloud platform it is detected
A type antibodies against foot-and-mouth disease virus standard curves, client is according to standard curve and nature controlling line and the fluorescence signal intensity value of detection line
Contrast relationship the potency of A types antibodies against foot-and-mouth disease virus in prepare liquid is calculated, according to potency result of determination.
7. application method according to claim 6, it is characterised in that：The standard curve is to import cloud in the following way
Platform, a collection of detection card, and A type antibodies against foot-and-mouth disease virus standard items corresponding to preparation are prepared first, block chromatography mark with detection
Quasi- product are simultaneously detected with chromatogram scanner, and the data processing that detection obtains is passed to client by chromatogram scanner, and client calculates
Detection line corresponding to standard items and the fluorescence signal intensity value of nature controlling line are drawn, and is returned according to this data so as to obtain A
Standard curve is transmitted to cloud platform and stored by type antibodies against foot-and-mouth disease virus standard curve, client.
8. application method according to claim 7, it is characterised in that：The standard curve that the client is calculated is formed
One file, and corresponding generation bar code, client can extract the standard curve by scanning bar code from cloud platform.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201711054389.8A CN107589269A (en) | 2017-11-01 | 2017-11-01 | Detection card for A type antibodies against foot-and-mouth disease virus in Quantitative detection serum |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201711054389.8A CN107589269A (en) | 2017-11-01 | 2017-11-01 | Detection card for A type antibodies against foot-and-mouth disease virus in Quantitative detection serum |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN107589269A true CN107589269A (en) | 2018-01-16 |
Family
ID=61044786
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201711054389.8A Pending CN107589269A (en) | 2017-11-01 | 2017-11-01 | Detection card for A type antibodies against foot-and-mouth disease virus in Quantitative detection serum |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN107589269A (en) |
Citations (12)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN204241487U (en) * | 2014-11-24 | 2015-04-01 | 河南科技学院 | The colloidal gold immune chromatography test card that a kind of Lomefloxacin and antibody thereof remain |
| CN204556641U (en) * | 2015-04-30 | 2015-08-12 | 苏州快捷康生物技术有限公司 | A kind of CDV immunochromatographydetection detection card |
| CN204594996U (en) * | 2015-04-29 | 2015-08-26 | 中国检验检疫科学研究院 | A kind of immune chromatography test paper holder |
| CN104964973A (en) * | 2015-07-08 | 2015-10-07 | 邓双胜 | Test paper reading and analyzing method and system based on mobile terminal camera |
| CN105606819A (en) * | 2016-01-21 | 2016-05-25 | 中国农业科学院兰州兽医研究所 | Detection card for serotype O foot and mouth disease virus antibody and preparation method of detection card |
| CN205353093U (en) * | 2016-02-04 | 2016-06-29 | 广州市微米生物科技有限公司 | Spacing formula immunofluorescence chromatography detects card |
| KR20160120675A (en) * | 2015-04-08 | 2016-10-18 | 주식회사 디에이치랩 | Rapid Quantitative Diagnostic Kit |
| CN106432434A (en) * | 2016-08-29 | 2017-02-22 | 中牧实业股份有限公司 | Enzyme-linked immunosorbent assay kit for detecting foot and mouth disease A-type structural protein VP1 antibody |
| CN206038693U (en) * | 2016-09-30 | 2017-03-22 | 三峡大学 | Immunofluorescence chromatography detection device |
| CN206038692U (en) * | 2016-09-28 | 2017-03-22 | 杭州华得森生物技术有限公司 | Modified colloidal gold detect reagent board |
| CN106680490A (en) * | 2016-12-03 | 2017-05-17 | 百奥森（江苏）食品安全科技有限公司 | Position limiting type immunofluorescence chromatography detection card |
| CN107037025A (en) * | 2017-06-24 | 2017-08-11 | 杭州微瑞科技有限公司 | The high quick fluorescent chromatographic device of group of the lanthanides and detection method |
-
2017
- 2017-11-01 CN CN201711054389.8A patent/CN107589269A/en active Pending
Patent Citations (12)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN204241487U (en) * | 2014-11-24 | 2015-04-01 | 河南科技学院 | The colloidal gold immune chromatography test card that a kind of Lomefloxacin and antibody thereof remain |
| KR20160120675A (en) * | 2015-04-08 | 2016-10-18 | 주식회사 디에이치랩 | Rapid Quantitative Diagnostic Kit |
| CN204594996U (en) * | 2015-04-29 | 2015-08-26 | 中国检验检疫科学研究院 | A kind of immune chromatography test paper holder |
| CN204556641U (en) * | 2015-04-30 | 2015-08-12 | 苏州快捷康生物技术有限公司 | A kind of CDV immunochromatographydetection detection card |
| CN104964973A (en) * | 2015-07-08 | 2015-10-07 | 邓双胜 | Test paper reading and analyzing method and system based on mobile terminal camera |
| CN105606819A (en) * | 2016-01-21 | 2016-05-25 | 中国农业科学院兰州兽医研究所 | Detection card for serotype O foot and mouth disease virus antibody and preparation method of detection card |
| CN205353093U (en) * | 2016-02-04 | 2016-06-29 | 广州市微米生物科技有限公司 | Spacing formula immunofluorescence chromatography detects card |
| CN106432434A (en) * | 2016-08-29 | 2017-02-22 | 中牧实业股份有限公司 | Enzyme-linked immunosorbent assay kit for detecting foot and mouth disease A-type structural protein VP1 antibody |
| CN206038692U (en) * | 2016-09-28 | 2017-03-22 | 杭州华得森生物技术有限公司 | Modified colloidal gold detect reagent board |
| CN206038693U (en) * | 2016-09-30 | 2017-03-22 | 三峡大学 | Immunofluorescence chromatography detection device |
| CN106680490A (en) * | 2016-12-03 | 2017-05-17 | 百奥森（江苏）食品安全科技有限公司 | Position limiting type immunofluorescence chromatography detection card |
| CN107037025A (en) * | 2017-06-24 | 2017-08-11 | 杭州微瑞科技有限公司 | The high quick fluorescent chromatographic device of group of the lanthanides and detection method |
Non-Patent Citations (2)
| Title |
|---|
| 张中旺，张永光: "抗原表位的研究方法及口蹄疫病毒抗原表位的研究进展", 《微生物学通报》 * |
| 高闪电 等: "A型口蹄疫病毒型特异性抗原的可溶性原核表达及多克隆抗体的制备", 《中国人兽共患病学报》 * |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Coleman et al. | Proteome and antigen profiling of Coxiella burnetii developmental forms | |
| CN101253261B (en) | Diagnostic test kits | |
| CN103760365B (en) | A kind of BAS-ELISA kit detecting CSFV Erns and E2 antigen | |
| CN105950563B (en) | The monoclonal antibody and application of hybridoma cell strain 7E3 and its resistant to foot and mouth disease A type virus of secretion | |
| WO2022032983A1 (en) | Quantum dot microsphere immunochromatography test strip for detection of african swine fever virus mucous membrane siga antibody, and use thereof | |
| CN107831317A (en) | Canine coronavirus antibody Quantitative detection card and application method | |
| CN107870236A (en) | Detection card for PPR antiviral antibody in Quantitative detection sheep blood serum | |
| CN103543261B (en) | Cattle and sheep Brucellosis indirect enzyme-linked immunosorbent assay antibody assay kit and preparation method thereof | |
| CN107884578A (en) | For echinococcosis antibody test card in Quantitative detection serum | |
| Ortiz et al. | Use of the 27-kilodalton recombinant protein from Paracoccidioides brasiliensis in serodiagnosis of paracoccidioidomycosis | |
| CN106432440A (en) | Brucella melitensis antibody PPA-ELISA detection kit and preparation method thereof | |
| CN102662063A (en) | Detection kit and method for non-structural protein antibody dot blots of foot and mouth disease viruses | |
| CN112858676A (en) | Fluorescent microsphere identification and detection card for African swine fever virus antibody | |
| CN107817352A (en) | Canine parvovirus antibody Quantitative detection card and application method | |
| CN104267181A (en) | African swine fever antigen and fluorescent nanocrystalline test strip prepared from African swine fever antigen | |
| CN107870243A (en) | For foot and mouth disease virus non-structural protein antibody test card in quick detection serum | |
| CN103820471A (en) | Recombined chlamydia trachomatis protein and application thereof | |
| CN107860921A (en) | Pseudorabies virus antibody Quantitative detection card and application method | |
| Lin et al. | Antibody responses of pigs to defined Erns fragments after infection with classical swine fever virus | |
| CN108333369A (en) | A kind of porcine reproductive and respiratory syndrome virus antibody detection method and its dedicated test card | |
| CN102533795B (en) | Recombinant human cytomegalovirus protein and applications thereof | |
| CN107589269A (en) | Detection card for A type antibodies against foot-and-mouth disease virus in Quantitative detection serum | |
| Van der Marel et al. | D-antigen determination in polio vaccine production: comparison of gel diffusion and ELISA methods. | |
| CN208432625U (en) | The rapid detection system of foot and mouth disease virus non-structural protein antibody | |
| CN208283402U (en) | The quantitative rapid detection system of PPR virus antibody |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| RJ01 | Rejection of invention patent application after publication | ||
| RJ01 | Rejection of invention patent application after publication |
Application publication date: 20180116 |