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Porcine epidemic diarrhea vaccine composition, and preparation method and application thereof

A porcine epidemic diarrhea and vaccine composition technology, which is applied in the direction of drug combinations, pharmaceutical formulas, medical preparations containing active ingredients, etc., can solve problems such as side effects, insufficient antigen content, and increased miscellaneous protein content

Active Publication Date: 2014-12-31
PU LIKE BIO ENG
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

There are potential biological risks in immunization with attenuated virus vaccines. The low titer of the whole virus inactivated vaccine leads to insufficient antigen content, and it is difficult to isolate and culture PEDV. Even if trypsin is added to the nutrient solution to promote its propagation in Vero cells, the virus titer Can only be maintained at 1×10 3.01 TCID 50 / mL or so
If the method of cell concentration is used to increase the content of miscellaneous proteins, causing side effects

Method used

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  • Porcine epidemic diarrhea vaccine composition, and preparation method and application thereof
  • Porcine epidemic diarrhea vaccine composition, and preparation method and application thereof
  • Porcine epidemic diarrhea vaccine composition, and preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0069] Example 1 Preparation, Identification and Content Determination of Porcine Epidemic Diarrhea Virus S1 Protein

[0070] 1.1 Preparation, identification and content determination of porcine epidemic diarrhea virus S1a protein

[0071] 1.1.1 Construction of recombinant vector Bacmid-S1a

[0072] According to the nucleotide sequence of porcine epidemic diarrhea virus S1 protein (see the sequence table SEQ ID No.1), it is synthesized by Shanghai Bioengineering Co., Ltd. by artificial synthesis, and the synthesized gene fragment has a full length of 2367bp. The template of S1 protein was prepared on the basis of the artificially synthesized S1 gene fragment.

[0073] The S1 full-length gene fragment synthesized in the previous step was used as a template for the PCR reaction, a pair of specific primers were designed, and XhoI and PstI restriction sites were added in the upstream and downstream respectively. The primer sequences are as follows:

[0074] S1a-P1: 5'GCTCTCGAGAT...

Embodiment 3

[0102] Example 3 Preparation of Porcine Epidemic Diarrhea Virus Vaccine Composition

[0103] 3.1 Preparation of porcine epidemic diarrhea virus subunit antigen

[0104] Dilute the recombinant proteins S1a, S1b, S1c, S1d, and S1e prepared in Example 1 using PBS at pH 7.2, and add aluminum gel adjuvant to mix well, so that the recombinant proteins S1a, S1b, S1c, and S1d in the vaccine composition The contents of S1e and S1e are shown in Table 1, while ensuring that the volume ratio of the aluminum gel adjuvant to the vaccine composition is 1:5. The prepared vaccine composition was used as an immunogen and stored at 4°C for future use.

[0105] 3.2 Preparation of porcine epidemic diarrhea whole virus inactivated antigen

[0106] The PEDV HN1301 strain virus solution was diluted with pH 7.2 PBS, and the diluted virus solution was inoculated with African green monkey kidney cells (Vero cells, purchased from Shanghai Institute of Biochemistry) covered with a dense monolayer by 3% ...

Embodiment 4

[0113] Example 4 Evaluation of Immunity Efficacy of Porcine Epidemic Diarrhea Vaccine Composition

[0114] 4.1 Active immunization test

[0115] Select 88 PEDV antibody-negative piglets at the age of 3 days, and randomly divide them into 11 groups (see Table 2 for details), 8 piglets / group, and the vaccine composition prepared in Example 3 of the first 1-1 to 1-9 groups are immunized successively through Houhai Point A-I, 1mL / head; groups 1-10 were immunized with the vaccine composition B prepared in Example 3 in Houhai Point; groups 1-11 were set as blank control groups, and were injected with the same dose of PBS solution at pH 7.2. On the 14th day after immunization, the sera of pigs in the immunized group and the control group were collected, and the titers of neutralizing antibodies in the sera of each group were detected. Statistical analysis was carried out on the average antibody titer of each piglet by using SPSS15.0 software. Simultaneously, the immune group (the 1...

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Abstract

The invention provides a porcine epidemic diarrhea vaccine composition. The vaccine composition contains a porcine epidemic diarrhea virus subunit antigen and a vector. The invention also provides a porcine epidemic diarrhea vaccine composition, and the vaccine composition comprises a porcine epidemic diarrhea virus subunit antigen, a porcine epidemic diarrhea totivirus inactivated antigen and a vector. The invention also discloses a preparation method of the vaccine composition and application thereof to preparation of drugs for preventing and / or treating porcine epidemic diarrhea. An S1 protein in the vaccine composition can conduct large amount of recombinant expression of components in the vaccine composition by means of genetic engineering, so as to realize short time consumption as well as facilitate large-scale production. When the vaccine composition contains the S1 protein and porcine epidemic diarrhea totivirus inactivated antigen, the vaccine composition has better immune effect than single usage of totivirus inactivated antigen, and can effectively prevent diseases caused by variants of porcine epidemic diarrhea and common strains.

Description

technical field [0001] The invention belongs to the technical field of veterinary biological products, and in particular relates to a porcine epidemic diarrhea vaccine composition and a preparation method and application thereof. Background technique [0002] Porcine epidemic diarrhea (porcine epidemic diarrhea, PED) is an important acute, contact, viral intestinal infectious disease caused by porcine epidemic diarrhea virus (porcine epidemic diarrhea virus, PEDV). Vomiting, dehydration, and decreased appetite are characteristic. The disease first occurred in the United Kingdom and Belgium in the 1970s. In 1973, PEDV was isolated in Shanghai, Liaoning, Jilin and other places in my country. PEDV can persist in pig herds, and pigs of all ages are susceptible. The morbidity rate of suckling piglets, shelf pigs and fattening pigs can reach 100%, especially severe with suckling piglets, and the case fatality rate reaches 100%. At present, there is no specific drug against PEDV...

Claims

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Application Information

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IPC IPC(8): A61K39/295A61K39/215A61P31/14A61P1/12
Inventor 张许科孙进忠王莹王延辉田克恭
Owner PU LIKE BIO ENG
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