CN104292300A - Epitope minimum motif peptide of P1, VP2 and VP4 structural proteins in type O foot and mouth disease virus (FMDV) strain (O/BY/CHA/2010) and application of epitope minimum motif peptide - Google Patents
Epitope minimum motif peptide of P1, VP2 and VP4 structural proteins in type O foot and mouth disease virus (FMDV) strain (O/BY/CHA/2010) and application of epitope minimum motif peptide Download PDFInfo
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Abstract
The invention belongs to the technical field of biological medicine and biological detection and particularly relates to an epitope minimum motif peptide of P1, VP2 and VP4 structural proteins in a type O foot and mouth disease virus (FMDV) strain (O/BY/CHA/2010) and application of the epitope. Five linear epitope minimum motif peptides provided by the invention can be used as candidate epitope peptides for research and development of a novel recombinant multi-epitope peptide vaccine against type O FMDV and have amino acid sequences as shown in SEQ. ID NO.1-SEQ. ID NO.5. The invention also provides a candidate epitope for research of a detection antigen of a high-specificity and high-sensitivity recombinant multi-epitope peptide for diagnosing prevalence of the FMDV and the candidate epitope has amino acid sequences as shown in the SEQ. ID NO.6-SEQ. ID NO.10.
Description
Technical field
The invention belongs to biological medicine and technical field of biological, be specifically related to the epitope minimum motif peptide of Structural protein VP1, VP2 and VP4 in O type foot and mouth disease virus (FMDV) strain (O/BY/CHA/2010), expansion peptide and application thereof.
Background technology
Livestock foot-and-mouth disease is the artiodactyls such as livestock contagious disease the most serious in the world today, main harm pig, ox, sheep, and foot and mouth disease is classified as first of category-A transmissible disease by World Organization for Animal Health.For many years, foot and mouth disease worldwide large-scale outbreak and popular, causes tremendous economic to lose to livestock industry.According to immunogenicity, foot and mouth disease virus has the hypotype of A, O, C, SAT I, SAT II, SAT III and Asia I totally 7 serotypes and more than 65.
Immunization is the main path of developing country's prevention foot and mouth disease outburst.Traditional deactivation vaccine has the advantages such as immunogenicity is high, protectiveness is good, but may occur vaccine inactivation not thoroughly or viral escape, thus causes the eruption and prevalence of foot and mouth disease, has potential danger.Therefore, Many researchers turns to the research to recombinant vaccine and multivalence peptide vaccine.With regard to the latter's recombinant multi-epitope peptide vaccine, desirable vaccine should comprise Linear B Cell Epitopes (hereinafter referred to as linear epitope) and t cell epitope simultaneously, the former brings out generation antibody in B cell, and the latter then can stimulate T cell to produce cytokine, impels B cell secretory antibody.Therefore, the Screening and Identification of both epi-positions is the important research content in virusology and vaccinology research field always, such as, just there is the research of relevant antigenic peptide to report (as Tang H before, Liu X S, Fang Y Z, et al.The Epitopes of Foot and Mouth Disease [J] .Asian Journal of Animal and Veterinary Advances, 2012,7 (12): 1261-1265; Blanco E, McCullough K, Summerfield A, et al.Interspecies major histocompatibility complex-restricted Th cell epitope on foot-and-mouth disease virus capsid protein VP4 [J] .J Virol, 2000,74:4902 – 4907; Wang Jialong etc.)。
But, although the authentication method of this respect hundreds and thousands of (Progress of Research Methods of Wu Jing and Wu Wutong .B cell protein epitope. Pharmaceutical Biotechnology, 7 (4): 239-242,2000; Shen Guanxin etc. translate. antibody technique experiment guide. and Science Press, 2002; The progress of the .B cell epitope location such as Wang Honglin. life science, 21:317-319,2009), as enumerated shown in document above, due to the restriction by linear (non-conformation type) B cell Epitope Identification method, what identified is all mostly be longer 16 poly-or longer antigenic peptides (antigenic peptide) without antibody recognition minimum motif (being usually made up of 3 ~ 8 residues) qualification result in the past, instead of real epitope peptide.In addition, (Shi YP under the background of recombinant multi-epitope peptide vaccine development is presented at synthetic peptide vaccine, Hasnain, SE, Sacci JB, et al.Immunogenicity and in vitro protective efficacy of a recombinant multistage Plasmodium falciparum candidate vaccine.Proc.Natl.Acad.Sci.USA.96,1615 – 162,1999; He YP, Xu WX, Hong AZ, et al.Immunogenic comparison for two different recombinant chimeric peptides (CP12 and CP22) containing one or two copies of three linear B cell epitopes from β-hCG subunit.J Biotechnol, 151:15-21,2011), in order to develop the effective preventative target virus multi-epitope peptide vaccine of antibody, the qualification of target protein linear epitope and antibody recognition minimum motif thereof just seems particularly important.
As everyone knows, from target protein identify one section can induce antibody generate antigenic peptide invention there is novelty and creativeness, therefore so both at home and abroad granted patent can be found everywhere.The meticulous epitope peptide that each antibody can identify is identified from target protein, also or its () be comprised in one section of published antigenic peptide sequence, it has novelty and creativeness equally, because it is the needs of practical application equally, as developed the occasion based on the recombinant multi-epitope peptide of epitope minimum motif peptide, just likely as much as possible combination linear epitope peptide (because for for being applicable to as intestinal bacteria and/or eukaryotic cell high expression level and development cost consideration, the artificial antigen peptide limited length of design, general about 200 amino-acid residues, wherein for guaranteeing that designed multi-epitope peptide has strong immunogenicity, the broad spectrum complementary and cytotoxic t cell epitope peptide for sars by force of general length 10 ~ 30 residues must be combined) simultaneously, also can contribute to producing specific required effective epitope antibodies (because Linear B Cell Epitopes is made up of maximum 8 residues of minimum 3 residues, just N number of epi-position may be there is in obvious long antigenic peptide, just likely produce N number of epitope antibodies, select meticulous epitope peptide that N number of epi-position number then can be made to reduce to minimum, in order to object antibody tormation, avoid other antibody tormation that may be harmful to) simultaneously.In a word, identify that the meaning of meticulous epitope peptide on invention target protein is apparent with application benefit.
It is to be noted, because foot and mouth disease is to the great effect of national economy, trade, have the title of " political economy is sick ", so long-standing for the research of foot and mouth disease virus vaccine, also be first successful identification epitope peptide and realize the virus (ref.) of gene chemical synthesis peptide vaccine, especially to study in the majority to O type FMDV.The people such as Pfaff as far back as VP 1 (144-159aa) LRGDLQVLAQKVARTL that nineteen eighty-two just identifies FMDV O 1K be a main antibody combining site (Pfaff E, Mussgay M,
hO, Schulz GE, Schaller H.Antibodies against a preselected peptide recognize and neutralize foot and mouth disease virus.EMBO J.1982, 1 (7): 869-74.), at same year, the people such as Bittle find FMDV O type, VP1 (25-41aa) (200-213) of Kaufbeuren strain is main B cell antigen epi-position (Bittle JL, Houghten RA, Alexander H, Shinnick TM, Sutcliffe JG, Lerner RA, Rowlands DJ, Brown F.Protection against foot-and-mouth disease by immunization with a chemically synthesized peptide predicted from the viral nucleotide sequence.Nature.1982 Jul 1, 298 (5869): 30-3.).But owing to being subject to the restriction of Epitope Identification method, be difficult to carry out minimum motif qualification.Because these 3 peptide sections do not have conservative property between FMDV is various or in type, the variation of FMDV strain is very fast in addition, once new epidemic situation occurs, must again set up specific aim vaccine.After this, the qualification about Asial I type coat protein epitope is also had, (1-12aa) of VP1, (17-29aa), (194-211aa); VP2 (40-50aa), VP3 (26-39aa), VP4 (30-41), but meticulous Epitope Identification (Zhang ZW is not carried out to it equally
1zhang YG, Wang YL, Pan L, Fang YZ, Jiang ST, L ü JL, Zhou P.Screening and identification of B cell epitopes of structural proteins of foot-and-mouth disease virus serotype Asia1.Vet Microbiol.2010 Jan6; 140 (1-2): 25-33.).More than study while identifying not a duck soup from the epitope scanning mapping of side reflection target protein and minimum motif, the epitope fine that also proved invents 5 O type FMDV identifies the demand of practical application really, can embody creativeness.
Summary of the invention
The object of the present invention is to provide the meticulous epitope peptide of 3 structural protein in foot and mouth disease virus (FMDV) O type strain (O/BY/CHA/2010), and the expansion peptide of these epitope minimum motif peptides (8 peptide) or be longer than the antigen of this small peptide (8 peptide); The expansion small peptide of described minimum motif peptide, minimum motif peptide (8 peptide) is also provided or is longer than the application of antigen of this small peptide (8 peptide).
Content of the present invention is more specifically described as follows.
In order to realize the object of the meticulous epitope peptide that the invention provides foot and mouth disease virus (FMDV) O type strain 3 structural protein, we are by improvement biosynthesizing peptide method, utilize cavy anti-FMDV O type standard serum, 3 structural protein of the FMDV O type strain O/BY/CHA/2010 of pop have carried out linear epitope scanning mapping research.Gather in peptide in the overlap 18 covering VP1, VP2 and VP4 tri-structural protein full length sequences, qualification can with the antigenic peptide of cavy anti-FMDV O type standard serum generation immunoblotting reaction; Overlapping 8 peptide sequences are designed further, by immunoblotting reaction and then determine its meticulous epitope and aminoacid sequence for antigenic peptide.Experimental result shows: the meticulous epitope peptide effectively having been identified 3 structural protein contained by foot and mouth disease virus O type strain (O/BY/CHA/2010) by improvement biosynthesizing peptide method and cavy anti-FMDV O type standard serum.
The invention provides the epitope minimum motif peptide of VP1, VP2 and VP4 tri-structural protein in foot and mouth disease virus O type strain (O/BY/CHA/2010), it is identified with cavy anti-FMDV O type standard serum, have 5 epi-positions, its aminoacid sequence is respectively: shown in SEQ.ID NO.1, SEQ.ID NO.2, SEQ.ID NO.3, SEQ.ID NO.4 and SEQ.ID NO.5.
Wherein, the aminoacid sequence of the minimum epitope motifs peptide of VP1 structural protein, for shown in SEQ.ID NO.1 and SEQ.ID NO.2, is designated as FMDV:VP1 successively
142-147, FMDV:VP1
204-208; The aminoacid sequence of the minimum epitope motifs peptide of VP2 structural protein, for shown in SEQ.ID NO.3 and SEQ.ID NO.4, is designated as FMDV:VP2 successively
8-13, FMDV:VP2
14-18; The aminoacid sequence of the minimum epitope motifs peptide of VP4 structural protein, for shown in SEQ.ID NO.5, is designated as FMDV:VP4
22-27.
The present invention also provides the expansion small peptide of above-mentioned 5 minimum epitope motifs peptides, and its aminoacid sequence is followed successively by shown in SEQ.ID No.6 ~ SEQ.ID No.10, and when chemosynthesis or amalgamation and expression albumen use, the residue moiety of expansion can additions and deletions or alternative.
The present invention also provides the described application of minimum epitope motifs peptide in the preparation of recombinant multi-epitope peptide vaccine, medicine or preparation detection antibody.
The present invention also provides the expansion small peptide of epitope minimum motif peptide alone or in combination in the application of the serum identification epitope tag thing of preparation foot and mouth disease epidemiological diagnosis.
The meticulous epitope peptide of above-mentioned foot and mouth disease virus O type strain (O/BY/CHA/2010) structural protein, also exist at other FMDV (O type, A type, Asial I type, C type, SAT I, SAT II, SAT III) epidemic isolates, namely build such recombinant multi-epitope peptide vaccine also effective to its alloytype foot and mouth disease virus strain.
Accompanying drawing explanation
Fig. 1 is the SDS-PAGE analytical results of 21 18 peptide amalgamation and expressions of VP1; The total protein that Figure 1A: swimming lane 1:BL21 (DE3) empty bacterium is expressed, the brachymemma GST188 albumen of swimming lane 2:pXXGST-1 plasmid expression, swimming lane 3 ~ 14: 18 poly-peptide P1 ~ P12 fusion roteins of overlapped 10 residues of covered structure albumen VP1 overall length of expression, swimming lane 15: molecular weight of albumen Marker; The total protein that Figure 1B: swimming lane 1:BL21 (DE3) empty bacterium is expressed, the brachymemma GST188 albumen of swimming lane 2:pXXGST-1 plasmid expression, swimming lane 3 ~ 11: the VP1 structural protein 18 of expression gather peptide P13 ~ P21 fusion rotein, swimming lane 12: molecular weight of albumen Marker.Object band is positioned at position corresponding to 25kDa place, and brachymemma GST188 carrier proteins is slightly less than restructuring target protein.In 1B figure swimming lane 5, object band is also 18 poly-peptides, and possible amino acid composition and primary structure difference cause mobility lower.
Fig. 2 is the SDS-PAGE analytical results of the 18 poly-peptide fusion proteins of expressing VP2 structural protein 26 overlapped 10 residues; The brachymemma GST188 albumen of Fig. 2 A swimming lane 1:pXXGST-1 plasmid expression, swimming lane 2 ~ 14: the VP2 structural protein 18 of expression gather peptide P1 ~ P13 fusion rotein, swimming lane 15: molecular weight of albumen Marker; The GST188 albumen of Fig. 2 B: swimming lane 1:pXXGST-1 plasmid expression, swimming lane 2 ~ 14: the VP2 structural protein 18 of expression gather peptide P14 ~ P26 fusion rotein, swimming lane 15: molecular weight of albumen Marker.Object band is positioned at position more on the lower side, 25kDa place, and brachymemma GST188 carrier proteins is slightly less than restructuring target protein.
Fig. 3 is the SDS-PAGE analytical results of expressing VP4 structural protein 10 18 poly-peptide fusion proteins: the GST188 albumen of swimming lane 1:pXXGST-1 plasmid expression, swimming lane 2 ~ 11: express VP4 structural protein 18 and gather peptide P1 ~ P10 fusion rotein, swimming lane 12: molecular weight of albumen Marker.Object band is positioned at position more on the lower side, 25kDa place, and brachymemma GST188 carrier proteins is slightly less than restructuring target protein.
Fig. 4 FMDV O/BY/CHA/2010 strain 3 structural protein 18 gather the immunoblotting qualification of peptide; Fig. 4 A: swimming lane 1 is the empty bacterium total protein contrast of BL21 (DE3), swimming lane 2 is pXXGST-1 expression GST188 protein control, and swimming lane 3 ~ 11 is followed successively by VP1 structural protein 18 and gathers peptide P13 ~ P21 fusion rotein, and swimming lane 12 is molecular weight of albumen Marker; Fig. 4 B: swimming lane 1 is pXXGST-1 plasmid expression GST188 protein control, and swimming lane 2 ~ 14 is followed successively by VP2 structural protein P1 ~ P13, and swimming lane 15 is molecular weight of albumen Marker; Fig. 4 C: swimming lane 1 is GST188 protein control, swimming lane 2 ~ 11 is followed successively by VP4 structural protein P1 ~ P10 fusion rotein, and swimming lane 12 is molecular weight of albumen Marker.
The 8 peptide clone construction and expression notes of Fig. 5 VP1 141aa-160aa: 1 is pXXGST-1 contrast, and 2 ~ 14 are followed successively by restructuring 8 peptide, and 15 is Marker.The protein band that object band is obvious between 25kDa ~ 15kDa, brachymemma GST188 carrier proteins is slightly less than restructuring target protein.
The 8 peptide clone construction and expressions of Fig. 6 VP1-14; Wherein: 1 is pXXGST-1 contrast, and 2 ~ 11 are followed successively by VP1-14-1 ~ VP1-14-10, and 12 is Marker.
The 8 peptide clone construction and expressions of Fig. 7 VP1-20; Wherein: 1 is pXXGST-1 contrast, and 2 ~ 12 are followed successively by VP1-20-1 ~ VP1-20-11, and 13 is Marker.
The 8 peptide clone construction and expressions of Fig. 8 VP1-21; Wherein: 1 is pXXGST-1 contrast, and 2 ~ 7 are followed successively by VP1-21-1 ~ VP1-21-6, and 8 is Marker.
The 8 peptide clone construction and expressions of Fig. 9 VP2-1; Wherein: 1 ~ 11 is followed successively by VP2-1-1 ~ VP2-1-11,12 is Marker.
The 8 peptide clone construction and expressions of Figure 10 VP2-2; Wherein: 1 is pXXGST-1 contrast, and 2 ~ 9 are followed successively by VP2-2-1 ~ VP2-2-8, and 10 is Marker.
The 8 peptide clone construction and expressions of Figure 11 VP4-3; Wherein: 1 is the empty bacterium contrast of BL21 (DE3), and 2 is pXXGST-1 contrast, and 3 ~ 13 are followed successively by VP4-3-1 ~ VP4-3-11, and 14 is Marker.
The 8 peptide immunoblotting qualifications of Figure 12 VP1 141-160; Wherein: 1 is pXXGST-1 contrast, and 2 ~ 14 are followed successively by VP1-N ~ VP1-Z, and 15 is Marker.
Figure 13 VP1-14, the 8 peptide immunoblotting qualifications of VP1-20, VP1-21; Wherein: Figure 13 A:1 is pXXGST-1 contrast, and 2 ~ 11 are followed successively by VP1-14-1 ~ VP1-14-10, and 12 is Marker; Figure 13 B:1 is pXXGST-1 contrast, and 2 ~ 12 are followed successively by VP1-20-1 ~ VP1-20-11, and 13 is Marker; Figure 13 C:1 is pXXGST-1 contrast, and 2 ~ 6 are followed successively by VP1-21-2 ~ VP1-21-6, and 7 is Marker.
The 8 peptide immunoblotting qualifications of Figure 14 VP2-1 and VP2-2; Wherein: Figure 14 A:1 is pXXGST-1 contrast, and 2 ~ 12 are followed successively by VP2-1-1 ~ VP2-1-11, and 13 is Marker; Figure 14 B:1 is pXXGST-1 contrast, and 2 ~ 9 are followed successively by VP2-2-1 ~ VP2-2-8, and 10 is Marker.
The 8 peptide immunoblotting qualifications of Figure 15 VP4-3; Wherein: 1 is pXXGST-1 contrast, and 2 ~ 13 are followed successively by VP4-3-1 ~ VP4-3-11, and 14 is Marker.
Embodiment
The present invention can describe further by following Examples.Illustrative example is not meaned and is limited scope involved in the present invention, and this scope is sufficiently clarified in description before.The experimental technique of unreceipted actual conditions in the following example, all conveniently condition and [U.S.] J. Pehanorm Brooker and D.W. Russell write the third edition " molecular cloning: the laboratory manual " (Science Press of the translations such as Huang Peitang, 2002) and [U.S.] E. breathe out " antibody technique experiment guide " (Science Press that Lip river and D. Lay grace write the translations such as Shen Guanxin, 2002) step described in, or carry out according to the condition of production and sales business suggestion.
Embodiment 1
The present embodiment is the qualification of FMDV O type strain O/BY/CHA/20104 the meticulous epitope of structural protein:
(1) the poly-peptide coding DNA fragment design of target protein series 18/8
According to the gene order (GenBank accession number JN998085.1) of VP1, VP2, VP4 tri-structural protein of FMDV O type strain O/BY/CHA/2010 in GenBank, after e. coli codon optimization is carried out to it, 18/8 poly-peptide coding DNA fragment of overlapped 10 the aa residues of design covering three protein gene full length sequences, and add BamH I and TAA-Sal I cohesive end sequence respectively at its two ends, the positive minus strand fragment of chemosynthesis each small peptide coding DNA.Complete the first round 18 gather pepscan mapping after, with regard to 8 poly-peptides of overlapped 7 residues of reactive peptide section design series, simultaneously for the poly-peptide of the epitope VP1 (21-40) found before and VP1 (141-160) design series 8, carry out second and take turns the qualification of antibody identification meter position minimum motif.
(2) structure of serial small peptide fusion protein expression plasmid, screening and qualification
By molecular cloning method, 18/8 of the synthesis poly-positive and negative chain encoding DNA fragmentation of peptide is annealed between two, then by DNA ligase they are connected with the carrier pXXGST-1 through BamH I and Sal I double digestion respectively and spend the night, product conversion E.coliBL21 (DE3) competence bacterial strain will be connected next day, be coated on and add on the LB flat board of Amp.Choosing single colony inoculation is added with in the LB liquid nutrient medium of Amp in 3mL, 220r/min, 30 DEG C shake bacterium 16 ~ 18h after, bacterium liquid is transferred to new 3mL in 2% ratio and is added with in the LB liquid nutrient medium of Amp, continue 220r/min, 30 DEG C cultivate 4h, until cell density OD
600light absorption value reaches 0.6, then in 42 DEG C of abduction delivering 5h; Get 1mL bacterium liquid collected by centrifugation thalline, add the ddH of 50 μ L
2after O suspension thalline, 2 × loading the Buffer adding 50 μ L again mixes, and places 5min in boiling water, the centrifugal 10min of 14000r/min, get 10 μ L supernatants and carry out SDS-PAGE electrophoresis (15% separation gel), negative control is for using vehicle Control pXXGST-1 transformed bacteria.Fig. 1-Fig. 3 is the SDS-PAGE result of 18 peptides; Fig. 5-Figure 11 is 8 peptide clone construction and expressions.
Get small peptide fusion rotein be greater than carrier proteins contrast 1 ~ 2kDa recombinant clone bacterium carry out DNA sequencing, verify the exactness of each synthetic peptide encode fragment sequence.
(3) antigen reactivity 18/8 gathers the immunoblotting qualification of peptide
The series 18/8 correct to DNA sequencing is gathered peptide clone bacterium and is carried out thermal induction expression, collect the immunoblotting qualification that total bacterial protein directly carries out, namely, electrophoresis is terminated each induction bacterium total protein electrotransfer on rear SDS-PAGE gel in the cellulose acetate membrane in 0.2 μm, aperture, 2h closed by the skimmed milk (preparing with PBST) with 5%; Add cavy anti-FMDV O type standard serum (skimmed milk with 5% is tired by it and diluted) and hatch 1h; 3 times are washed, 5min/ time with PBST; The anti-cavy IgG of rabbit (skimmed milk with 5% is tired by it and diluted) adding horseradish peroxidase-labeled hatches 1h; 3 times are washed, 5min/ time with washings; With exposure instrument, experimental result is detected (with ECL substrate luminescence reagent box).Fig. 4 is the immunoblotting qualification result of 18 poly-peptides; Figure 12-Figure 15 is the immunoblotting qualification result of 8 peptide clones.
(4) determination of antigen epitope minimum motif
It is as shown in table 1 that the reactivity 8 screened gathers peptide,
The 18 poly-peptides that table 1 immunoblotting reaction screens and the determination of 8 poly-peptides and meticulous epi-position
Consensus sequence according to them determines meticulous epitope VP1 142-147 (PNVRGD), VP1204-208 (KIVAP), VP2 8-13 (TLLEDR), VP2 14-18 (ILTTR), VP4 22-27 (INNYYM).
Claims (4)
1. the epitope minimum motif peptide of middle VP1, VP2 and VP4 tri-structural protein of foot and mouth disease virus O type strain (O/BY/CHA/2010), have 5 epi-positions, its aminoacid sequence is respectively shown in SEQ.ID NO.1, SEQ.ID NO.2, SEQ.ID NO.3, SEQ.ID NO.4 and SEQ.ID NO.5.
2. an expansion small peptide for minimum epitope motifs peptide as claimed in claim 1, is characterized in that aminoacid sequence is followed successively by shown in SEQ.ID No.6 ~ SEQ.ID No.10; When chemosynthesis or amalgamation and expression albumen use, the residue moiety of expansion can additions and deletions or alternative.
3. the minimum epitope motifs peptide of middle VP1, VP2 and VP4 tri-structural protein of foot and mouth disease virus O type strain (O/BY/CHA/2010) as claimed in claim 1, detects the application of antibody aspect in the preparation of recombinant multi-epitope peptide vaccine, medicine or preparation.
4. the expansion small peptide of epitope minimum motif peptide as claimed in claim 2 is in the application alone or in combination in the serum identification epitope tag thing of preparation foot and mouth disease epidemiological diagnosis.
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| CN108931644A (en) * | 2018-07-19 | 2018-12-04 | 河南省农业科学院 | A kind of evaluation of foot and mouth disease virus immune antiboidy and infection diagnose bigeminy test strips with Immune dctection |
| CN108761093B (en) * | 2018-07-19 | 2020-12-25 | 河南百奥生物工程有限公司 | Test strip for evaluating foot-and-mouth disease virus antibody |
| CN108931644B (en) * | 2018-07-19 | 2021-09-10 | 河南省农业科学院 | Foot-and-mouth disease virus immune antibody evaluation and infection and immune differential diagnosis dual test strip |
| CN117304277A (en) * | 2023-09-26 | 2023-12-29 | 中国农业科学院兰州兽医研究所 | O-type foot-and-mouth disease virus VP4 protein T cell epitope polypeptide and application thereof |
| CN117304277B (en) * | 2023-09-26 | 2024-03-08 | 中国农业科学院兰州兽医研究所 | O-type foot-and-mouth disease virus VP4 protein T cell epitope polypeptide and application thereof |
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