CN108761069A - A kind of newcastle disease virus immune colloid gold Rapid detection test strip and preparation method thereof established using epitope antibodies - Google Patents
A kind of newcastle disease virus immune colloid gold Rapid detection test strip and preparation method thereof established using epitope antibodies Download PDFInfo
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- CN108761069A CN108761069A CN201810308275.XA CN201810308275A CN108761069A CN 108761069 A CN108761069 A CN 108761069A CN 201810308275 A CN201810308275 A CN 201810308275A CN 108761069 A CN108761069 A CN 108761069A
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- G—PHYSICS
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- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/558—Immunoassay; Biospecific binding assay; Materials therefor using diffusion or migration of antigen or antibody
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/569—Immunoassay; Biospecific binding assay; Materials therefor for microorganisms, e.g. protozoa, bacteria, viruses
- G01N33/56983—Viruses
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Abstract
The newcastle disease virus immune colloid gold Rapid detection test strip and preparation method thereof that the invention discloses a kind of to establish using epitope antibodies.Technical scheme of the present invention main points are：The bottom of test strips is the supporting layer that PVC board is constituted, sticking successively on the supporting layer has closely coupled sample pad, gold labeling antibody bonding pad, nitrocellulose filter and absorption pad, the polypeptide P25 epitope antibodies of colloid gold label are wherein adsorbed on gold labeling antibody bonding pad, the nature controlling line of the detection line and the printing of rabbit anti-chicken IgG solution of the printing of newcastle disease virus mAb solution is coated on nitrocellulose filter, the amino acid sequence of polypeptide P25 is TCPDEQDYQLRMA (Thr Cys Pro Asp Glu Gln Asp Tyr Gln Leu Arg Met Ala).The invention further particularly discloses the preparation methods of the newcastle disease virus immune colloid gold Rapid detection test strip.The newcastle disease virus immune colloid gold Rapid detection test strip of the present invention detects quick and high specificity, convenient for quickly detection NDV is viral at the scene.
Description
Technical field
The invention belongs to the preparing technical fields of immuno-chromatographic test paper strip, and in particular to it is a kind of using epitope antibodies establish
Newcastle disease virus immune colloid gold Rapid detection test strip and preparation method thereof.
Background technology
Newcastle disease (Newcastle Disease) is by newcastle disease virus (Newcastle disease virus, NDV)
Caused one kind is acute, hot, septic and high tactile property birds infectious disease, and incidence and mortality is all very high, seriously endangers
The development of China's aviculture.Newcastle disease virus belongs to paramyxovirus section (Paramyxoviridae), fowl Rubulavirus,
Sub-thread minus-stranded rna virus has cyst membrane, full-length genome 15186bp to encode 6 structural proteins, including phosphoprotein (P), core
Albumen (NP), stromatin (M), six hatching egg of fusion protein (F), large protein (L) and hemagglutinin-neuraminidase protein (HN)
In vain.Hemagglutinin-neuraminidase albumen (HN) has hemagglutinin activity (HA) and nervonic acid ammonia enzymatic activity (NA) and can cooperate with F eggs
It plays a role in vain, in virus infection, identifies host cell surface sialic acid receptor to HN protein-specifics, this is virus
Necessary to infecting host cell, and body can be induced to generate protective immunity antibody.
Currently, common detection method has hemagglutinin reaction (hemagglutinin, HA), hemagglutination inhibition reaction
(hemagglutinin inhibition, HI), enzyme-linked immunosorbent assay (enzyme-linked immunosorbent
Assay, ELISA) and the detection methods such as real-time quantitative PCR, however these methods all compare time and effort consuming, need laboratory special
Equipment could detect, and cannot diagnosis be made to popular propagate of newcastle disease virus in time.Glue is immunized in existing NDV
Body gold detection method is to prepare NDV monoclonal antibodies by murine myeloma cell, then marks colloidal gold to prepare NDV and quickly examines
Paper slip is tested, the method needs to prepare the cumbersome processes such as the antigen of purifying, immune mouse and raising zooblast.
Invention content
The technical problem to be solved by the present invention is to provide a kind of newcastle disease virus immune colloid gold Rapid detection test strips
And preparation method thereof, 13 polypeptides are synthesized according to newcastle disease virus HN protein designs, it is anti-with newcastle disease virus clinic respectively
Body carries out ELISA detections, and screening polypeptide P25 (its amino acid sequence is TCPDEQDYQLRMA) can be with newcastle disease virus
Specific reaction occurs, polypeptide P25 is adsorbed in affinity column, screening is for polypeptide P25 from Hyper Immunized Serum on Chicken Newcastle Disease
Epitope antibodies, then will polypeptide P25 epitope antibodies mark colloidal gold after prepare Rapid detection test strip for quickly detection chicken it is new
City epidemic disease poison.
The present invention adopts the following technical scheme that newcastle disease virus immune colloid gold is quickly examined to solve above-mentioned technical problem
Test paper slip, it is characterised in that：The bottom of the test strips is the supporting layer that PVC board is constituted, and sticking successively on the supporting layer has
Closely coupled sample pad, gold labeling antibody bonding pad, nitrocellulose filter and absorption pad, is wherein adsorbed on gold labeling antibody bonding pad
There are the polypeptide P25 epitope antibodies of colloid gold label, the detection of newcastle disease virus mAb solution printing is coated on nitrocellulose filter
The nature controlling line of line and the printing of rabbit anti-chicken IgG solution, the distance between detection line and nature controlling line are 5mm, the amino acid sequence of polypeptide P25
It is classified as TCPDEQDYQLRMA.
Further preferably, the width that laminates of the sample pad and gold labeling antibody bonding pad is 2mm, gold labeling antibody bonding pad
The width that laminates with nitrocellulose filter is 2mm, and the width that laminates of absorption pad and nitrocellulose filter is 2mm.
The preparation method of newcastle disease virus immune colloid gold Rapid detection test strip of the present invention, it is characterised in that
The specific steps are：
(1) preparation of newcastle disease virus hyper-immune serum
Newcastle disease virus is inoculated in chicken fibroblasts, newcastle disease virus is harvested after 72h, with the side of sucrose gradient centrifugation
Method purifies newcastle disease virus, and the chicken of 2 week old is given with the concentration intramuscular injection of 1mg/mL, blood is acquired when there are clinical symptoms
Clearly, antibody titer is detected by ELISA method with polypeptide P25, and the serum being collected into is inactivated into 30min in 56 DEG C, in -70 DEG C
It saves backup；
(2) preparation of polypeptide P25 epitope antibodies
According toPolypeptide P25 is diluted to 500 μ g/mL and marked by Immobilization Kit kits to exempt from
Epidemic disease affinity column finally uses elution by the newcastle disease virus hyper-immune serum of preparation by immune affinity chromatographic column
Polypeptide P25 epitope antibodies repeat 2-3 times, the polypeptide P25 epitope antibodies being collected into are saved backup in -20 DEG C；
(3) preparation of gold labeling antibody bonding pad
Polypeptide P25 epitope antibodies are removed into sediment through 15000rpm centrifugations 30min in 4 DEG C, are added successively in centrifuge tube
Enter colloidal gold solution and polypeptide P25 epitope antibodies, is protected from light stands 30min at room temperature, add 1/10 colloidal gold solution volume
20mmol/L sodium tetraborate solution containing 10wt%BSA centrifuges 30min through 13000rpm in 4 DEG C, discards supernatant, use and colloid
Colloidal gold is resuspended in the isometric 20mmol/L sodium tetraborate solution containing 2wt%BSA of gold solution, is centrifuged through 13000rpm in 4 DEG C
30min is discarded supernatant, with the 20mmol/L sodium tetraborate solution suspension colloids containing 1wt%BSA of 1/2 colloidal gold solution volume
Gold completes the work of colloid gold label polypeptide P25 epitope antibodies, is placed in 4 DEG C and saves backup；
(4) preparation of nitrocellulose filter
Newcastle disease virus mAb and rabbit anti-chicken IgG are diluted to 1mg/mL by 1 μ L/cm even applications to nitric acid with PBS respectively
It is used as detection line and nature controlling line, detection line and nature controlling line to be located at the center of nitrocellulose filter, room temperature at a distance of 5mm on cellulose membrane
It spontaneously dries, is saved backup in 4 DEG C after encapsulation；
(5) assembling of test strips
Colloid gold label polypeptide P25 epitope antibodies are adsorbed on bonding pad and obtain gold labeling antibody bonding pad, then by sample
Pad, gold labeling antibody bonding pad, nitrocellulose filter and absorption pad affix to PVC board structure after keeping the overlapping widths of 2mm in order
At supporting layer on, be then cut into the strip test strips that width is 4mm, then test strips be fitted into test card, be put into baking
Kept dry is spare in oven.
The present invention has the advantages that compared with prior art：Using the polypeptide P25 epitope antibodies of colloid gold label
It is successfully prepared ndv antigen Rapid detection test strip, which shows higher sensitivity and spy
The opposite sex, it is highly consistent with hemagglutination experimental result with other poultry diseases such as IBDV and AIV no cross reactions；With HA
Experiment is compared, and NDV Rapid detection test strips can detect 5U NDV, and NDV can be detected in 3-5min, and not depend on
In Other Instruments and reagent, therefore convenient for quickly detection NDV is viral at the scene.
Description of the drawings
Fig. 1 is the structural schematic diagram of newcastle disease virus immune colloid gold Rapid detection test strip of the present invention；
Fig. 2 is the detects schematic diagram of newcastle disease virus immune colloid gold Rapid detection test strip of the present invention；
Fig. 3 is newcastle disease virus immune colloid gold Rapid detection test strip sensitivity detection figure of the present invention；
Fig. 4 is newcastle disease virus immune colloid gold Rapid detection test strip specific detection figure of the present invention.
Specific implementation mode
The above of the present invention is described in further details by the following examples, but this should not be interpreted as to this
The range for inventing above-mentioned theme is only limitted to embodiment below, and all technologies realized based on the above of the present invention belong to this hair
Bright range.
Embodiment
As shown in Figure 1, newcastle disease virus immune colloid gold Rapid detection test strip, the bottom of the test strips is PVC
The supporting layer that plate is constituted, sticking successively on the supporting layer has closely coupled sample pad, gold labeling antibody bonding pad, nitrocellulose
Film and absorption pad are wherein adsorbed with the polypeptide P25 epitope antibodies of colloid gold label, nitrocellulose filter on gold labeling antibody bonding pad
On be coated with the printing of newcastle disease virus mAb solution detection line and the printing of rabbit anti-chicken IgG solution nature controlling line, detection line and Quality Control
The distance between line is 5mm, and the amino acid sequence of polypeptide P25 is TCPDEQDYQLRMA.
The preparation method of newcastle disease virus immune colloid gold Rapid detection test strip is：
(1) preparation of newcastle disease virus hyper-immune serum
Newcastle disease virus is inoculated in chicken fibroblasts, newcastle disease virus is harvested after 72h, with the side of sucrose gradient centrifugation
Method purifies newcastle disease virus, and the chicken of 2 week old is given with the concentration intramuscular injection of 1mg/mL, blood is acquired when there are clinical symptoms
Clearly, antibody titer is detected by ELISA method with polypeptide P25, and the serum being collected into is inactivated into 30min in 56 DEG C, in -70 DEG C
It saves backup；
(2) preparation of polypeptide P25 epitope antibodies
According toPolypeptide P25 is diluted to 500 μ g/mL and marked by Immobilization Kit kits to exempt from
Epidemic disease affinity column finally uses elution by the newcastle disease virus hyper-immune serum of preparation by immune affinity chromatographic column
Polypeptide P25 epitope antibodies repeat 2-3 times, the polypeptide P25 epitope antibodies being collected into are saved backup in -20 DEG C；
(3) preparation of gold labeling antibody bonding pad
Polypeptide P25 epitope antibodies are removed into sediment through 15000rpm centrifugations 30min in 4 DEG C, are added successively in centrifuge tube
Enter colloidal gold solution and polypeptide P25 epitope antibodies, is protected from light stands 30min at room temperature, add 1/10 colloidal gold solution volume
20mmol/L sodium tetraborate solution containing 10wt%BSA centrifuges 30min through 13000rpm in 4 DEG C, discards supernatant, use and colloid
Colloidal gold is resuspended in the isometric 20mmol/L sodium tetraborate solution containing 2wt%BSA of gold solution, is centrifuged through 13000rpm in 4 DEG C
30min is discarded supernatant, with the 20mmol/L sodium tetraborate solution suspension colloids containing 1wt%BSA of 1/2 colloidal gold solution volume
Gold completes the work of colloid gold label polypeptide P25 epitope antibodies, is placed in 4 DEG C and saves backup；
(4) preparation of nitrocellulose filter
Newcastle disease virus mAb and rabbit anti-chicken IgG are diluted to 1mg/mL by 1 μ L/cm even applications to nitric acid with PBS respectively
It is used as detection line and nature controlling line, detection line and nature controlling line to be located at the center of nitrocellulose filter, room temperature at a distance of 5mm on cellulose membrane
It spontaneously dries, is saved backup in 4 DEG C after encapsulation；
(5) assembling of test strips
Colloid gold label polypeptide P25 epitope antibodies are adsorbed on bonding pad and obtain gold labeling antibody bonding pad, then by sample
Pad, gold labeling antibody bonding pad, nitrocellulose filter and absorption pad affix to PVC board structure after keeping the overlapping widths of 2mm in order
At supporting layer on, be then cut into the strip test strips that width is 4mm, then test strips be fitted into test card, be put into baking
Kept dry is spare in oven.
Immune chromatography test paper testing principle with the blood serum sample to be checked of normal saline dilution as shown in Fig. 2, drop to sample pad
After upper, under capillary action, flowed from sample pad one end toward absorption pad one end, if containing NDV in sample, gold labeling antibody can be therewith
In conjunction with Ag-Ab-colloidal gold composite is formed, the compound of formation is when being moved at detection line and nature controlling line, in sample
NDV to be combined with the newcastle disease virus mAb in detection line and form a red line be detection line (T lines), the rabbit on nature controlling line
Anti-chicken IgG will then be combined with the gold labeling antibody of polypeptide P25, and to form another red line be nature controlling line (C lines).Due to negative NDV
Without containing NDV virus, therefore the newcastle disease virus mAb in detection line cannot with Ag-Ab-colloidal gold composite in conjunction with and block
Gold labeling antibody is cut, so without band at detection line, but the rabbit anti-chicken IgG on nature controlling line but can be anti-with polypeptide P25 gold marks always
Body in conjunction with and form a red line, therefore can judge whether test strips effective by nature controlling line.
Hemagglutination test detects NDV viruses
It can be aggregated chicken red blood cell according to all newcastle disease virus strains, and be macroscopic, hemagglutination examination
Test the goldstandard that detection NDV virions are newcastle disease detections.25 μ L PBS are added in V-Bottom microwell plate respectively first, so
The test sample of 25 μ L is placed in the first hole afterwards, NDV is diluted to 2 successively-1-2-9.The 1% of 25 μ L is added into each hole
Then red blood cell gently beats the side 15min of plate at room temperature, read and record the result in each hole.
After room temperature 15min, red blood cell is aggregated completely, the results showed that 2-8It is red thin that diluted virus liquid can be aggregated chicken completely
Born of the same parents.Therefore, the HA potency of NDV strains is 2-8, i.e., it is 2 that the hemagglmination of NDV strains, which concentrates the 1U of virus titer,-8。
The sensitivity and specificity of test strips detect
The virus titer of the sample of NDV strains is measured with hemagglutination test (HA), then NDV test strips is used to examine
Survey the serial dilution degree containing NDV viruses 1-50U.1U in the virus titer of hemagglutinin detection refers to cause
The virus for the minimum that red blood cell is aggregated completely.Meanwhile feeling infectious bursa of Fabricius virus (IBDV) and avian influenza virus (AIV) use
Make negative control.
Ndv antigen test strip from different batches detects the sensitivity of NDV viruses respectively, compared with haemagglutination,
Ndv antigen Rapid detection test strip is able to detect that 5U NDV (Fig. 3).By being detected to 2 positive NDV samples, IBDV
Strain and AIV strains are used as negative control.The result shows that 2 NDV samples occur red item in p-wire and control line
Band, and only there is 1 band (Fig. 4) at control line in negative control.
Clinical sample detects
Collection different regions, which are suspected to be, infects 86 parts of NDV virosis chicken tissues, including tracheae, cecal tonsil, buccal swab,
Intestines etc., take a little tissue sample respectively, are added 1mL PBS grindings, and after multigelation 3 times, 12000rpm centrifuges 1min.It uses respectively
HA and the detection of ndv antigen test strip.
Ndv antigen Rapid detection test strip and HA is used to test 86 NDV samples (table 1) from different regions respectively.Knot
Fruit shows：Ndv antigen Rapid detection test strip and HA testing results are highly consistent, rate of accuracy reached to 98.8% (85/86).
1 NDV clinical detection results of table
Newcastle disease is one of the epidemic disease for endangering China's aviculture most serious, is listed as with highly pathogenic bird flu universally acknowledged
Most important 2 birds epidemic diseases.The disease is a kind of animal epidemic disease as defined in the disease that must be reported as defined in OIE and China
The anti-system of newcastle disease and research are attached great importance to always in disease, countries in the world.HN albumen is a kind of important multifunctional meter of NDV viruses
Face glycoprotein is made of cytoplasmic domain, transmembrane region, stem area and ball heads, wherein there are receptor binding domain, neuraminic acids
Enzyme active sites and epitope.Pre-stage test shows that the peptide P25 of newcastle disease virus HN albumen can be with NDV hyper-immuneserums
Reaction.The method of affinity chromatography is utilized to screen the epitope antibodies for polypeptide P25 from NDV hyper-immuneserums in the present invention.
The present invention is successfully prepared ndv antigen Rapid detection test strip using the polypeptide P25 epitope antibodies of colloid gold label,
The ndv antigen Rapid detection test strip shows higher sensitivity and specificity, with other poultry diseases such as IBDV and AIV without
Cross reactivity, it is highly consistent with hemagglutination experimental result；Compared with HA is tested, NDV Rapid detection test strips can be examined
5U NDV are surveyed, and NDV can be detected in 3-5min, and independent of Other Instruments and reagent, therefore convenient at the scene
Quickly detection NDV viruses.
Embodiment above describes the basic principles and main features and advantage of the present invention, and the technical staff of the industry should
Understand, the present invention is not limited to the above embodiments, and the above embodiments and description only describe the originals of the present invention
Reason, under the range for not departing from the principle of the invention, various changes and improvements may be made to the invention, these changes and improvements are each fallen within
In the scope of protection of the invention.
Sequence table
<110>Xinxiang University
<120>A kind of newcastle disease virus immune colloid gold Rapid detection test strip and its preparation using epitope antibodies foundation
Method
<160> 1
<170> SIPOSequenceListing 1.0
<210> 1
<211> 13
<212> PRT
<213>Artificial sequence (Artificial sequence)
<400> 1
Thr Cys Pro Asp Glu Gln Asp Tyr Gln Leu Arg Met Ala
1 5 10
Claims (3)
1. a kind of newcastle disease virus immune colloid gold Rapid detection test strip established using epitope antibodies, it is characterised in that：
The bottom of the test strips is the supporting layer that PVC board is constituted, and sticking successively on the supporting layer has closely coupled sample pad, Jin Biao
Antibody bonding pad, nitrocellulose filter and absorption pad are wherein adsorbed with the polypeptide P25 of colloid gold label on gold labeling antibody bonding pad
Epitope antibodies are coated with detection line and the printing of rabbit anti-chicken IgG solution of the printing of newcastle disease virus mAb solution on nitrocellulose filter
Nature controlling line, the distance between detection line and nature controlling line are 5mm, and the amino acid sequence of polypeptide P25 is TCPDEQDYQLRMA (Thr
Cys Pro Asp Glu Gln Asp Tyr Gln Leu Arg Met Ala)。
2. a kind of newcastle disease virus immune colloid gold established using epitope antibodies according to claim 1 quickly detects
Test strips, it is characterised in that：The width that laminates of the sample pad and gold labeling antibody bonding pad is 2mm, gold labeling antibody bonding pad
The width that laminates with nitrocellulose filter is 2mm, and the width that laminates of absorption pad and nitrocellulose filter is 2mm.
3. a kind of a kind of newcastle disease virus immune colloid gold using epitope antibodies foundation described in claim 1 is quickly examined
Test the preparation method of paper slip, it is characterised in that the specific steps are：
（1）The preparation of newcastle disease virus hyper-immune serum
Newcastle disease virus is inoculated in chicken fibroblasts, newcastle disease virus is harvested after 72h, it is pure with the method for sucrose gradient centrifugation
Change newcastle disease virus, the chicken of 2 week old is given with the concentration intramuscular injection of 1mg/mL, serum is acquired when there are clinical symptoms, uses
Polypeptide P25 detects antibody titer by ELISA method, and the serum being collected into is inactivated 30min in 56 DEG C, in -70 DEG C of preservations
It is spare；
（2）The preparation of polypeptide P25 epitope antibodies
Polypeptide P25 is diluted to 500 μ g/mL and labelled immune according to SulfoLink Immobilization Kit kits
Affinity column is finally more with elution by the newcastle disease virus hyper-immune serum of preparation by immune affinity chromatographic column
Peptide P25 epitope antibodies repeat 2-3 times, the polypeptide P25 epitope antibodies being collected into are saved backup in -20 DEG C；
（3）The preparation of gold labeling antibody bonding pad
Polypeptide P25 epitope antibodies are removed into sediment through 15000rpm centrifugations 30min in 4 DEG C, glue is sequentially added in centrifuge tube
Body gold solution and polypeptide P25 epitope antibodies are protected from light stand 30min at room temperature, add containing for 1/10 colloidal gold solution volume
The 20mmol/L sodium tetraborate solution of 10wt% BSA centrifuges 30min through 13000rpm in 4 DEG C, discards supernatant, use and colloidal gold
Colloidal gold is resuspended in the 20mmol/L sodium tetraborate solution of the isometric BSA containing 2wt% of solution, is centrifuged through 13000rpm in 4 DEG C
30min is discarded supernatant, with the 20mmol/L sodium tetraborate solution suspension colloids of the BSA containing 1wt% of 1/2 colloidal gold solution volume
Gold completes the work of colloid gold label polypeptide P25 epitope antibodies, is placed in 4 DEG C and saves backup；
（4）The preparation of nitrocellulose filter
Newcastle disease virus mAb and rabbit anti-chicken IgG are diluted to 1mg/mL by 1 μ L/cm even applications to cellulose nitrate with PBS respectively
It is used as detection line and nature controlling line, detection line and nature controlling line to be located at the center of nitrocellulose filter at a distance of 5mm on plain film, room temperature is natural
It is dry, it is saved backup in 4 DEG C after encapsulation；
（5）The assembling of test strips
Colloid gold label polypeptide P25 epitope antibodies are adsorbed on bonding pad and obtain gold labeling antibody bonding pad, then by sample pad, gold
Labeling antibody bonding pad, nitrocellulose filter and absorption pad keep affixing to after the overlapping widths of 2mm in order the branch of PVC board composition
It supports on layer, is then cut into width and is the strip test strips of 4mm, then test strips are fitted into test card, be put into baking box
Kept dry is spare.
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| CN201810308275.XA CN108761069A (en) | 2018-04-09 | 2018-04-09 | A kind of newcastle disease virus immune colloid gold Rapid detection test strip and preparation method thereof established using epitope antibodies |
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111551744A (en) * | 2020-05-15 | 2020-08-18 | 安徽中起生物科技有限公司 | Newcastle disease virus N protein IgY antibody colloidal carbon detection test paper and application thereof |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN104459142A (en) * | 2014-12-12 | 2015-03-25 | 河南省农业科学院 | Test paper for distinguishing virulent strain from attenuated strain of newcastle disease virus |
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| Title |
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| 于幼平 主编: "《鸡传染性法氏囊病的诊断与防治》", 31 January 1993, 北京农业大学出版社 * |
| 孙佳善: "禽流感、新城疫和犬痘热病毒单克隆抗体筛选策略比较及应用研究", 《中国博士学位论文全文数据库（农业科技辑）》 * |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN111551744A (en) * | 2020-05-15 | 2020-08-18 | 安徽中起生物科技有限公司 | Newcastle disease virus N protein IgY antibody colloidal carbon detection test paper and application thereof |
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Application publication date: 20181106 |