CN109836478B - Preparation method and application of African swine fever virus P11.5 protein specific polyclonal antibody - Google Patents
Preparation method and application of African swine fever virus P11.5 protein specific polyclonal antibody Download PDFInfo
- Publication number
- CN109836478B CN109836478B CN201910208025.3A CN201910208025A CN109836478B CN 109836478 B CN109836478 B CN 109836478B CN 201910208025 A CN201910208025 A CN 201910208025A CN 109836478 B CN109836478 B CN 109836478B
- Authority
- CN
- China
- Prior art keywords
- swine fever
- african swine
- fever virus
- polyclonal antibody
- protein
- 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.)
- Active
Links
Landscapes
- Peptides Or Proteins (AREA)
Abstract
The invention discloses a preparation method and application of a P11.5 protein specific polyclonal antibody of African swine fever virus, and the method utilizes P11.5 polypeptide epitope to immunize experimental animals such as mice, rabbits and the like to prepare the specific polyclonal antibody. The method has the advantages of high antigen purity, strong specificity of the prepared antibody, high affinity, small amount of used antigen and simple and convenient operation. The preparation method of the polyclonal antibody comprises the steps of selection of specific epitope of ASFV P11.5, synthesis of polypeptide antigen, animal immunization, determination of antibody and the like. Compared with the conventional method, the method has the advantages of simplicity, convenience, rapidness, high antigen purity, strong specificity of the prepared antibody and small antigen consumption. The polyclonal antibody can be used for detecting the protein P11.5 of the African swine fever virus, and provides an important technical method for prevention and control of the African swine fever in China.
Description
Technical Field
The invention relates to a preparation method and application of an African swine fever virus P11.5 protein specific polyclonal antibody.
Background
African swine fever is also known as east non-swine fever or wart swine disease, and is an acute heat type highly-contagious disease of pigs caused by African swine fever virus. The disease only infects pigs, and the clinical symptoms and pathological changes are similar to swine fever, mainly manifested as fever, cyanosis of skin, bleeding of lymph nodes, kidney, gastrointestinal mucosa and the like. The mortality rate of the disease is very high and can reach 100 percent during the circulation. At present, the African swine fever is already epidemic in more than 30 countries, and has a tendency of continuously expanding and spreading, and the first African swine fever epidemic situation is diagnosed in China in 8 months and 3 days in 2018. The diagnosis of the African swine fever is usually carried out by adopting an erythrocyte adsorption test, a direct immunofluorescence test, an animal inoculation test, an indirect immunofluorescence test, an enzyme-linked immunosorbent test, an immunoelectrophoresis test, an indirect enzyme-linked immunosorbent assay and the like, and the related specific reagents in the world are lack, so that a plurality of difficulties are brought to the control of the African swine fever. The African swine fever virus is double-stranded DNA containing an envelope, the genome is linear double-stranded DNA, the length is between 170 and 190kb, and 151 proteins can be encoded. The virus is the only member of African swine fever virus family, and at present, only 1 serotype exists, but the genomes of ASFV isolates in different regions have certain differences. According to genetic evolution analysis of the P72 gene, ASFV can be currently divided into 23 genotypes.
Disclosure of Invention
In order to overcome the defects, the invention provides a preparation method and application of a polyclonal antibody specific to the African swine fever virus P11.5 protein. The P11.5 gene is relatively stable and is very abundant in infected cells, and the invention utilizes the expression of the P11.5 gene, polypeptide synthesis and the like to establish a specific antibody and an antigen for rapidly detecting the African swine fever so as to provide a high-quality reagent for detecting the African swine fever.
In order to achieve the purpose of the invention, the invention adopts the technical scheme that: an African swine fever virus P11.5 protein specific antigen polypeptide, wherein the antigen polypeptide is P11.5-2 or P11.5-1; the sequence of P11.5-2 is PEERCTYKFNSYTKKMEL, and the sequence of P11.5-1 is DQEEKKALQNKETKNLGIP.
The antigen polypeptide is prepared by synthesis.
The invention also provides an African swine fever virus P11.5 protein immunogen which is directly composed of the African swine fever virus P11.5 protein specific antigen polypeptide; or the immunogen is formed by mixing the African swine fever virus P11.5 protein specific antigen polypeptide and a complete adjuvant; or the immunogen is formed by mixing the African swine fever virus P11.5 protein specific antigen polypeptide and an incomplete adjuvant.
The invention also discloses a preparation method of the African swine fever virus P11.5 protein specific polyclonal antibody, which comprises the following steps:
a) selecting a polypeptide epitope specific to the African swine fever virus P11.5 protein:
P11.5-1—DQEEKKALQNKETKNLGIP；
or P11.5-2-PEERCTYKFNSYTKKMEL;
b) synthesizing a specific polypeptide epitope of the African swine fever virus P11.5 protein;
c) firstly, mixing the African swine fever virus P11.5 protein specific polypeptide epitope with a complete adjuvant or an incomplete adjuvant and then immunizing animals; then, directly immunizing animals with the African swine fever virus P11.5 protein specific polypeptide epitope;
d) separating serum of the immunized animal, wherein the serum contains an anti-African swine fever virus P11.5 protein specific polyclonal antibody.
Preferably, the animal is a mouse or a rabbit.
The invention also provides an anti-African swine fever virus P11.5 protein specific polyclonal antibody prepared by the method and application of the African swine fever virus P11.5 protein specific polyclonal antibody in preparation of a kit for detecting an African swine fever virus antigen and/or antibody.
Compared with the prior art, the invention has the beneficial effects that: the invention discloses a method for quickly preparing a specific polyclonal antibody of African swine fever virus P11.5 protein, which utilizes P11.5 polypeptide epitope to immunize experimental animals such as mice, rabbits and the like to prepare the specific polyclonal antibody. The method has the advantages of high antigen purity, strong specificity of the prepared antibody, high affinity, small antigen consumption and simple operation. The preparation method of the polyclonal antibody comprises the steps of selection of specific epitope of ASFV P11.5, synthesis of polypeptide antigen, animal immunization, determination of antibody and the like. Compared with the conventional method, the method has the advantages of simplicity, convenience, rapidness, high antigen purity, strong specificity of the prepared antibody and small antigen consumption. The polyclonal antibody can be used for detecting the protein P11.5 of the African swine fever virus, and provides an important technical method for prevention and control of the African swine fever in China.
Detailed Description
The invention is further illustrated below with reference to specific examples. It should be understood that these examples are for illustrative purposes only and are not intended to limit the scope of the present invention. Further, it should be understood that various changes or modifications can be made to the present invention by those skilled in the art after reading the teaching of the present invention, and these equivalents also fall within the scope of the appended claims.
1. Screening of polypeptide epitopes
According to the amino acid sequence of the African swine fever virus P11.5 protein disclosed on line in medicine, a specific epitope with good hydrophilicity, strong antigenicity and commonality is screened and designed, such as the sequences P11.5-1-DQEEKKALQNKETKNLGIP (SEQ ID No.1) and P11.5-2-PEERCTYKFNSYTKKMEL (SEQ ID No. 2).
2. Synthesis of polypeptide epitopes
Synthesizing linear polypeptide, and synthesizing the polypeptide from C end to N end.
1. 3g of RINK resin (degree of substitution 0.3mmol/g) was weighed into a 150ml reactor and soaked with 50ml of Dichloromethane (DCM).
After 2.2 hours, the resin was washed with 3 resin volumes of N-Dimethylformamide (DMF) and then drained, and this was repeated four times and the resin was drained until use.
3. The Fmoc protecting group on the resin was removed by adding a quantity of 20% piperidine (piperidine/DMF) to the reactor and shaking on a decolourising shaker for 20 min. After deprotection, the resin was washed four times with 3 resin volumes of DMF and then drained.
4. And (3) detecting a small amount of resin by a ninhydrin (ninhydrin hydrate nine-well) method (detecting two drops of the resin A and B respectively, and reacting at 100 ℃ for 1min), wherein the resin has a color, which indicates that deprotection is successful.
5. Weighing a proper amount of the first amino acid at the C end and a proper amount of 1-hydroxy-benzotriazole (HOBT) into a 50ml centrifuge tube, adding 20ml of DMF to dissolve the DMF, then adding 3ml of N, N-Diisopropylcarbodiimide (DIC) to shake and shake for 1min, adding the solution into a reactor after the solution is clarified, and then placing the reactor into a 30 ℃ shaking table to react.
After 6.2 hours, the reaction mixture was capped with a suitable amount of acetic anhydride (acetic anhydride: DIEA: DCM ═ 1: 1: 2) for half an hour, washed four times with 3 resin volumes of DMF and drained.
7. The Fmoc protecting group on the resin was removed by adding a quantity of 20% piperidine (piperidine/DMF ═ 1:4) to the reactor and shaking on a decolourizing shaker for 20 min. After deprotection was performed four times with DMF and then drained.
8. And (3) detecting a small amount of resin by a ninhydrin (ninhydrin) method (detecting A and B, respectively, and reacting at 100 ℃ for 1min), wherein the resin is colored, which indicates that the deprotection is successful.
9. Weighing a second proper amount of amino acid and a proper amount of HOBT in a 50ml centrifuge tube, adding 25ml of DMF to dissolve the amino acid and the HOBT, adding 2.5ml of DIC to shake and shake for 1min, adding the solution into a reactor after the solution is clarified, and then placing the reactor in a shaking table at 30 ℃ to react.
After 10.1 hours, a small amount of resin is taken for detection, and the detection is carried out by an indanthrone method (two drops are respectively detected A and B, and the reaction is carried out for 1min at 100 ℃), if the resin is colorless, the reaction is complete; if the resin is colored, the condensation is not complete and the reaction is continued.
11. After the reaction was complete, the resin was washed four times with DMF and then drained, and a quantity of 20% piperidine (piperidine/DMF ═ 1:4) was added to the reactor and shaken on a decolorizing shaker for 20min to remove the Fmoc protecting group from the resin. After the protection is removed, washing with DMF for four times, and then draining to detect whether the protection is removed.
12. The following amino acids are sequentially ligated according to steps 9-11.
13. After the last amino acid had been grafted, the protection was removed, washed four times with DMF and the resin was drained with methanol. The polypeptide was then cleaved from the resin with 95 cleavage medium (trifluoroacetic acid: 1,2 ethanedithiol: 3, isopropylsilane: water: 95:2:2:1) (10 ml of cleavage medium per gram of resin) and centrifuged four times with glacial ethyl ether (cleavage medium: ethyl ether: 1: 9). And finally, separating and purifying by using HPLC (high performance liquid chromatography), and freeze-drying to obtain the polypeptide with certain purity.
Wherein HPLC separation and purification conditions are as follows:
stationary phase: c18;
configuration of mobile phase:
pump A: v (TFA)/V (Water) 1/1000
Pump B v (tfa)/v (acetonitrile) 1/1000;
flow rate: 10 mL/min;
retention time: 20-30 min.
3. Immunization of polypeptides
a. Mixing polypeptide antigen (P11.5-1 or P11.5-2) with complete adjuvant, and immunizing mouse or intracutaneous and plantar rabbit via intraperitoneal route; the immunization dose is 5ug per mouse, and the immunization dose of the rabbit is 10ug per mouse. b. Mixing the polypeptide antigen with incomplete adjuvant 7 days later, and immunizing mice or rabbits intradermally, intraplantarly and intranodal; the immunization dose is 10ug per mouse, and the immunization dose of the rabbit is 50ug per mouse.
c. 7 days later, the polypeptide antigen is directly used for immunizing mice or rabbits in the skin and lymph nodes by an abdominal cavity route; the immunization dose is 20ug per mouse, and the immunization dose of the rabbit is 100ug per mouse.
d. Blood was collected 7 days later, and serum was separated to determine antibody reactivity. If the titer is insufficient, the immunization can be repeated once more.
e. The separated serum contains polyclonal antibody specific to the P11.5 protein of African swine fever virus. And (3) coating an enzyme label plate by using hemocyanin-coupled polypeptide, performing ELISA detection, and after serum is diluted by 1:100, wherein the reactivity of an antibody is more than 1.0 of OD450 or OD 490.
3. Use of polyclonal antibodies
1) ELISA (enzyme-Linked immuno sorbent assay) detection of African swine fever virus P11.5 protein specific polyclonal antibody
The enzyme label plate is coated by a synthesized polypeptide antigen (P11.5-2), other non-specific active groups are sealed by a sealing liquid, after washing, the anti-African swine fever virus P11.5 protein specific polyclonal antibody serum of a mouse to be detected is added, after incubation for 1 hour at 37 ℃, washing is carried out, then the enzyme label anti-mouse IgG whole molecular antibody is incubated for 1 hour at 37 ℃, a substrate is used for developing color, and whether the African swine fever virus P11.5 protein specific polyclonal antibody exists is judged according to the developing intensity. The results are shown in tables 1 and 2.
TABLE 1 detection of African swine fever virus P11.5 protein-specific polyclonal antibody (P11.5-1) by ELISA method
TABLE 2 detection of African swine fever virus P11.5 protein-specific polyclonal antibody (P11.5-2) by ELISA method
As can be seen from the above results, the P11.5-2 polypeptide had better antigenicity than P11.5-1. The P11.5-2 polypeptide antigen was therefore selected. Wherein 1:100 means mouse serum was measured as 1:100 dilution. The data in the table are the readings for ELISA, the higher the reading, the higher the antibody titer.
If the swine serum is detected to contain the polyclonal antibody specific to the African swine fever virus P11.5 protein, the same method and principle can be adopted, and the technical personnel can completely understand and operate the method as long as the swine serum is replaced by the swine serum, and the enzyme-labeled antibody is replaced by the anti-swine IgG and/or IgM enzyme-labeled antibody.
2) Or detecting the African swine fever virus P11.5 protein specific polyclonal antibody by a blocking ELISA method, coating an enzyme label plate by using a synthesized polypeptide antigen (P11.5-2), sealing other non-specific active groups by using a sealing liquid, adding rabbit (or pig) anti-African swine fever virus P11.5 protein specific polyclonal antibody serum to be detected after washing, incubating for 30 minutes at 37 ℃, washing, adding a mouse anti-African swine fever virus P11.5 protein specific polyclonal antibody, incubating for 30 minutes at 37 ℃, incubating for 1 hour at 37 ℃ with an enzyme-labeled anti-mouse IgG whole molecular antibody after washing, developing color by using a substrate, calculating the OD value inhibition rate according to the developed color intensity, judging whether the African swine fever virus P11.5 protein specific polyclonal antibody exists, and judging the antibody is positive if the OD value inhibition rate reaches 35% or more. As in table 3 below:
TABLE 3 detection of African swine fever virus P11.5 protein specific polyclonal antibody by blocking ELISA method
| Negative control | Blank control | Rabbit serum 1 | Rabbit serum 2 | Rabbit serum 3 | Rabbit serum 4 | |
| P11.5-2 | 1.150 | 0.06 | 0.312 | 0.223 | 0.212 | 1.10 |
| Inhibition rate | 72.87％ | 80.61％ | 81.56％ | 4.34％ | ||
| Results | Positive for | Positive for | Positive for | Negative of |
3) Sandwich ELISA for detecting African swine fever virus P11.5 specific protein
The ELISA plate is coated with polyclonal antibody against P11.5 (rabbit serum obtained by P11.5-2 immunization, purified IgG), unbound nonspecific group is sealed by confining liquid, a sample to be detected (expressed African swine fever virus P11.5 protein or tissue sample of pig) is added, incubation is carried out at 37 ℃ for 1 hour, washing is carried out, enzyme-labeled anti-P11.5 mouse IgG enzyme-labeled antibody (mouse serum obtained by P11.5-2 or P11.5-1 immunization, purified IgG) is added, incubation is carried out at 37 ℃ for 1 hour, and substrate color development is carried out after washing. According to the intensity of the color development, whether the African swine fever P11.5 protein antigen exists in the sample is judged, and the result is shown in a table 4.
TABLE 4 Sandwich ELISA assay for African swine fever virus P11.5 specific protein antigen results
| Expression product | Blank control | Negative control | |
| OD650 value | 1.203 | 0.075 | 0.066 |
| Results | Positive for | Negative of | Negative of |
4) Other applications of polyclonal antibodies, such as immunofluorescence assays and the like
Firstly, constructing a pcDNA-P11.5 vector for expressing African swine fever virus P11.5 protein, transfecting vero cells, after 48 hours, incubating with the polyclonal antibody serum (serum obtained by P11.5-2 immunization), washing after 30 minutes, incubating with fluorescein FITC labeled anti-mouse IgG whole molecule antibody for 30 minutes, washing, adding 50% glycerol PBS buffer solution for mounting, and observing under a fluorescence microscope to obtain specific bright green fluorescence.
The examples described are illustrative of the invention and are not to be construed as limiting the invention, and any variations and modifications which come within the meaning and range of equivalency of the invention are to be considered within the scope of the invention.
Sequence listing
<110> Yangzhou university
Preparation method and application of <120> African swine fever virus P11.5 protein specific polyclonal antibody
<130> xhx2019031901
<141> 2019-03-19
<160> 2
<170> SIPOSequenceListing 1.0
<210> 1
<211> 19
<212> PRT
<213> Reticuloendotheliosis virus
<400> 1
Asp Gln Glu Glu Lys Lys Ala Leu Gln Asn Lys Glu Thr Lys Asn Leu
1 5 10 15
Gly Ile Pro
<210> 2
<211> 18
<212> PRT
<213> Reticuloendotheliosis virus
<400> 2
Pro Glu Glu Arg Cys Thr Tyr Lys Phe Asn Ser Tyr Thr Lys Lys Met
1 5 10 15
Glu Leu
Claims (6)
1. An African swine fever virus P11.5 protein specific antigen polypeptide, which is characterized in that the antigen polypeptide is P11.5-2 or P11.5-1; the sequence of P11.5-2 is PEERCTYKFNSYTKKMEL, and the sequence of P11.5-1 is DQEEKKALQNKETKNLGIP.
2. An African swine fever virus P11.5 protein immunogen, characterized in that, the immunogen is directly composed of the African swine fever virus P11.5 protein specific antigen polypeptide of claim 1; or, the immunogen is formed by mixing the African swine fever virus P11.5 protein specific antigen polypeptide of claim 1 and a complete adjuvant; alternatively, the immunogen is prepared by mixing the African swine fever virus P11.5 protein specific antigen polypeptide of claim 1 and an incomplete adjuvant.
3. A preparation method of an African swine fever virus P11.5 protein specific polyclonal antibody comprises the following steps:
a) selecting a polypeptide epitope specific to the African swine fever virus P11.5 protein: P11.5-1-DQEEKKALQNKETKNLGIP;
or P11.5-2-PEERCTYKFNSYTKKMEL;
b) synthesizing a specific polypeptide epitope of the African swine fever virus P11.5 protein;
c) firstly, mixing the African swine fever virus P11.5 protein specific polypeptide epitope with a complete adjuvant or an incomplete adjuvant and then immunizing animals; then directly immunizing animals with the African swine fever virus P11.5 protein specific polypeptide epitope;
d) separating serum of the immunized animal, wherein the serum contains an anti-African swine fever virus P11.5 protein specific polyclonal antibody.
4. The method of claim 3, wherein the animal is a mouse or a rabbit.
5. An anti-African swine fever virus P11.5 protein specific polyclonal antibody prepared according to the method of claim 3.
6. The African swine fever virus P11.5 protein specific polyclonal antibody of claim 5, in the preparation of a kit for detecting African swine fever virus antigen.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201910208025.3A CN109836478B (en) | 2019-03-19 | 2019-03-19 | Preparation method and application of African swine fever virus P11.5 protein specific polyclonal antibody |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201910208025.3A CN109836478B (en) | 2019-03-19 | 2019-03-19 | Preparation method and application of African swine fever virus P11.5 protein specific polyclonal antibody |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN109836478A CN109836478A (en) | 2019-06-04 |
| CN109836478B true CN109836478B (en) | 2022-06-14 |
Family
ID=66885911
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201910208025.3A Active CN109836478B (en) | 2019-03-19 | 2019-03-19 | Preparation method and application of African swine fever virus P11.5 protein specific polyclonal antibody |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN109836478B (en) |
Families Citing this family (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN112209992B (en) * | 2020-08-26 | 2021-12-17 | 中国农业科学院兰州兽医研究所 | Polypeptide for promoting swine organisms to generate African swine fever virus antigen specific immune response and application thereof |
Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO1993025666A1 (en) * | 1992-06-16 | 1993-12-23 | Commonwealth Scientific And Industrial Research Organisation | Recombinant entomopoxvirus |
| CN1840698A (en) * | 2006-01-06 | 2006-10-04 | 云南出入境检验检疫局检验检疫技术中心 | Fluorescence quantitative PCR detection reagent for Asf virus and preparation method and use thereof |
| CN101068934A (en) * | 2003-10-29 | 2007-11-07 | 里伯米德生物技术公司 | Compositions, methods and detection technologies for reiterative oligonucleotide synthesis |
| CN102066415A (en) * | 2007-10-29 | 2011-05-18 | 弗吉尼亚科技知识产权有限公司 | Porcine DC-SIGN, ICAM-3 and LSECtin and uses thereof |
| CN104267181A (en) * | 2014-10-17 | 2015-01-07 | 深圳出入境检验检疫局动植物检验检疫技术中心 | African swine fever antigen and fluorescent nanocrystalline test strip prepared from African swine fever antigen |
| US9474797B1 (en) * | 2014-06-19 | 2016-10-25 | The United States Of America, As Represented By The Secretary Of Agriculture | African swine fever virus georgia strain adapted to efficiently grow in the vero cell line |
-
2019
- 2019-03-19 CN CN201910208025.3A patent/CN109836478B/en active Active
Patent Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO1993025666A1 (en) * | 1992-06-16 | 1993-12-23 | Commonwealth Scientific And Industrial Research Organisation | Recombinant entomopoxvirus |
| CN101068934A (en) * | 2003-10-29 | 2007-11-07 | 里伯米德生物技术公司 | Compositions, methods and detection technologies for reiterative oligonucleotide synthesis |
| CN1840698A (en) * | 2006-01-06 | 2006-10-04 | 云南出入境检验检疫局检验检疫技术中心 | Fluorescence quantitative PCR detection reagent for Asf virus and preparation method and use thereof |
| CN102066415A (en) * | 2007-10-29 | 2011-05-18 | 弗吉尼亚科技知识产权有限公司 | Porcine DC-SIGN, ICAM-3 and LSECtin and uses thereof |
| US9474797B1 (en) * | 2014-06-19 | 2016-10-25 | The United States Of America, As Represented By The Secretary Of Agriculture | African swine fever virus georgia strain adapted to efficiently grow in the vero cell line |
| CN104267181A (en) * | 2014-10-17 | 2015-01-07 | 深圳出入境检验检疫局动植物检验检疫技术中心 | African swine fever antigen and fluorescent nanocrystalline test strip prepared from African swine fever antigen |
Also Published As
| Publication number | Publication date |
|---|---|
| CN109836478A (en) | 2019-06-04 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| US5595915A (en) | Method of determining antigenically active amino acid sequences | |
| Gomara et al. | Synthetic peptides for the immunodiagnosis of human diseases | |
| CN111647053A (en) | Polypeptide and application thereof in novel coronavirus detection and antibody or vaccine screening | |
| JP3188717B2 (en) | Hepatitis C assay | |
| CN100429234C (en) | Compounds and methods of early diagnosis of cervical cancer and genital condyloma with hpv, chsp60 tumor suppressor h-ras, k-ras and pten derived peptides modified | |
| EP0138854A1 (en) | Antigenically active amino acid sequences. | |
| RU2158928C2 (en) | Method and reagents for identifying hepatitis c virus | |
| WO1984003506A1 (en) | Antigenically active amino acid sequences | |
| CN108473540B (en) | Mutant HEV polypeptides and their use for determining anti-HEV antibodies | |
| KR930004055B1 (en) | Peptide and its use | |
| CA2391356A1 (en) | Peptides designed for the diagnosis and treatment of rheumatoid arthritis | |
| EP1328811B1 (en) | Hcv mosaic antigen composition | |
| Chang et al. | Development and comparison of enzyme-linked immunosorbent assays based on recombinant trimeric full-length and truncated spike proteins for detecting antibodies against porcine epidemic diarrhea virus | |
| WO2012134416A2 (en) | Peptide ligands | |
| US5824506A (en) | Dengue virus peptides and methods | |
| CN109836478B (en) | Preparation method and application of African swine fever virus P11.5 protein specific polyclonal antibody | |
| Ma et al. | Varying abilities of recombinant polypeptides from different regions of hepatitis E virus ORF2 and ORF3 to detect anti‐HEV immunoglobulin M | |
| JP2000514300A (en) | Peptide reagent for detecting human cytomegalovirus (CMV) | |
| US20050100883A1 (en) | Peptide-based diagnostic reagents for SARS | |
| US20140106381A1 (en) | Method for the diagnosis of rheumatoid arthritis | |
| US20060263765A1 (en) | Peptides and mixtures thereof for use in the detection of severe acute respiratory syndrome-associated coronavirus (sars) | |
| Pál et al. | Sandwich type ELISA and a fluorescent cytometric microbead assay for quantitative determination of hepatitis B virus X antigen level in human sera | |
| CN114324874A (en) | African swine fever virus detection kit and application thereof | |
| CN108948174B (en) | Citrulline modified peptide and application thereof | |
| CN113444154A (en) | Polypeptide and application thereof in novel coronavirus detection and antibody or vaccine screening |
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 | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |