Porcine pseudorabies virus strain, inactivated vaccine prepared from porcine pseudorabies virus strain and application of porcine pseudorabies virus strain

A technology for porcine pseudorabies virus and porcine pseudorabies, applied in antiviral agents, viruses/bacteriophages, biochemical equipment and methods, etc., can solve problems such as differences in immunogenicity, achieve effective prevention and control, good safety, and good mental state Effect

Active Publication Date: 2015-03-04
哈药集团生物疫苗有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Due to the continuous mutation of the epidemic strains, there are differences in immunogenicity between the commercially available vaccines and the vaccines prepared from traditional strains, which cannot prevent and control the current epidemic of porcine pseudorabies.

Method used

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  • Porcine pseudorabies virus strain, inactivated vaccine prepared from porcine pseudorabies virus strain and application of porcine pseudorabies virus strain
  • Porcine pseudorabies virus strain, inactivated vaccine prepared from porcine pseudorabies virus strain and application of porcine pseudorabies virus strain
  • Porcine pseudorabies virus strain, inactivated vaccine prepared from porcine pseudorabies virus strain and application of porcine pseudorabies virus strain

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] The isolation of embodiment 1 porcine pseudorabies virus strain

[0031] 1. Experimental method

[0032] 1.1 Disease data collection

[0033] The disease materials were collected from the brain tissue and tonsil of stillborn sows in a diseased pig herd in Jilin. Add DMEM at a ratio of 1:10, grind, prepare tissue suspension, freeze and thaw three times, centrifuge at 12,000rpm for 10min, and take the supernatant through a 0.22μm filter filter. The filtrate was stored in a -80°C refrigerator.

[0034] 1.2 Isolation and culture

[0035] The above-mentioned disease material was inserted into the Vero cell culture of the green monkey kidney cell line that had not yet formed a monolayer according to the content of 10% of the virus culture solution, placed at 37 ° C for 1 hour, and DMEM culture solution containing 2% calf serum was added, Cultured at 37°C for 5 days. After 2 times of freezing and thawing, the second generation was cultured for 4 days, and the culture medi...

Embodiment 2

[0042] The identification of embodiment 2 porcine pseudorabies virus strains

[0043] The PRV-JL strain of porcine pseudorabies virus isolated in Example 1 of the present invention was identified.

[0044] 1. Virus plaque cloning

[0045] The isolated porcine pseudorabies virus PRV-JL strain was diluted 10 times with maintenance solution, and 10 of them were taken. -5 、10 -6 、10 -7 、10 -8 Inoculate 24-well cell plates, inoculate 4 wells for each dilution, 100 μl / well. After incubating in a 37°C incubator for 1h, add 1mL of 44°C nutrient agar to each well, turn the cell plate over after the nutrient agar solidifies, and place in a place containing 5% CO 2 In the cell culture incubator, observe daily. After 4 days of culture, count the plaques at each dilution, calculate the plaque forming units (PFU), observe the shape and size of the plaques, pick out small and isolated plaques, add them to 0.5mL nutrient solution, freeze and thaw for 2 Once, 3,000r / min, centrifuge for ...

experiment example 1

[0061] Preparation of Experimental Example 1 Porcine Pseudorabies Inactivated Vaccine

[0062] 1. Preparation of antigen solution for seedling production

[0063] The PRV-JL strain of porcine pseudorabies virus isolated in Example 1 is inserted into the Vero cell culture forming a monolayer by 1% of the virus maintenance liquid volume, and placed in a 37°C rotary culture. When the lesion reaches 80%, the harvest contains the virus. Cell culture medium, after 2 times of freezing and thawing, collected the poison.

[0064] 2. Inactivation

[0065] According to 0.02% (g / mL) of the total amount of virus liquid, add 2% (g / mL) divinyl imine solution to the virus liquid, shake it fully, inactivate it on a shaker at 30°C and 100r / min for 60h, then add 2% sodium thiosulfate solution, terminate the inactivation, and test for sterility.

[0066] 3. Concentration

[0067] The inactivated virus liquid was taken, centrifuged horizontally at 4,000r / min, and then filtered with a 0.2um mem...

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Abstract

The invention discloses a porcine pseudorabies virus strain, an inactivated vaccine prepared from the porcine pseudorabies virus strain and an application of the porcine pseudorabies virus strain and belongs to the fields of separating and applying the porcine pseudorabies virus strain. The invention firstly discloses a porcine pseudorabies virus PRV-JL strain, and a microorganism preservation number of the porcine pseudorabies virus PRV-JL strain is CGMCC No.9903. The invention also discloses a method of applying the PRV-JL strain to preparing the porcine pseudorabies inactivated vaccine. The method comprises the following steps: (1) amplifying the porcine pseudorabies virus PRV-JL strain, and harvesting a virus liquid; (2) adding an inactivator to inactivate the virus liquid, and concentrating; (3) preparing a water phase and an oil phase; and (4) mixing the water phase and the oil phase, and emulsifying to obtain the porcine pseudorabies inactivated vaccine. Proved by animal experiment results that the inactivated vaccine prepared by the PRV-JL strain is high in safety, is good in immunogenicity, is high in immune protective rate, is capable of providing complete protection for both newly separated variant strain virulent strains and PRV MIN A strain virulent strains and is capable of effectively controlling current porcine pseudorabies.

Description

technical field [0001] The present invention relates to porcine pseudorabies virus strains, in particular to a virulent strain of porcine pseudorabies virus (Pseudorabies Virus) with excellent performance. The invention belongs to the field of isolation and application of porcine pseudorabies virus strains. Background technique [0002] Porcine pseudorabies is caused by porcine pseudorabies virus and often spreads in outbreaks. The death rate of newborn piglets infected with the virus is extremely high, and the adult pigs infected with the virus are mostly in a recessive state, and the death rate is low, but pregnant sows are prone to miscarriage, weak fetuses, dead and mummified fetuses. Healthy pigs can be infected with the virus by direct contact with infected pigs and sick pigs. The immunosuppressive effect of the virus increases the susceptibility of infected adult pigs to other pathogens, resulting in increased mortality in virus-infected pigs. In recent years, the ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N7/00A61K39/245A61P31/22C12R1/93
Inventor 王全杰张智明丁国杰步帆王彬窦海燕杨朋欣李来旭张凤强张祎冯闻迪
Owner 哈药集团生物疫苗有限公司
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