Duck astrovirus vaccine and preparation method thereof

An astrovirus and vaccine technology, applied in biochemical equipment and methods, vaccines, viruses, etc., can solve the problems of no disease, economic losses in the duck industry, etc., and achieve good immunity, easy concentration processing, and huge application prospects. Effect

Active Publication Date: 2019-04-30
广东渔跃生物技术有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] At present, there is no duck astrovirus vaccine on the market, and there is no related drug to contr...

Method used

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  • Duck astrovirus vaccine and preparation method thereof
  • Duck astrovirus vaccine and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0025] Preparation:

[0026] (1) Digest and disperse BHK-21 cells with trypsin, add 5% newborn bovine serum in DMEM culture solution in a spinner bottle at 37°C, 5% CO 2 After culturing to a monolayer of cells, the original culture medium was discarded, and the cells were washed 3 times with serum-free DMEM medium for inoculation of duck astrovirus type Ⅰ and type Ⅲ.

[0027] (2) Virus inoculation and cultivation: respectively inoculate the duck astrovirus type Ⅰ and type Ⅲ viruses into the cells prepared in step ① according to the final volume of 1:100, and absorb the virus solution at 37°C for 30 minutes, discard the virus liquid, and DMEM medium containing 1% newborn bovine serum at 37°C, 5% CO 2 Culture until the cells are terminated when lesions appear.

[0028] (3) Virus collection, concentration and purification and determination of virus content: the collected cell virus liquid was repeatedly frozen and thawed twice at -20°C, centrifuged at 4°C and 5000rpm for 10min,...

Embodiment 2

[0032] Preparation:

[0033] (1) Digest and disperse the BHK-21 cells with trypsin, add 10% newborn calf serum to the DMEM culture solution in a microcarrier reactor at 37°C and 5% CO 2 After culturing to a monolayer of cells, the original culture medium was discarded, and the cells were washed 3 times with serum-free DMEM medium for inoculation of duck astrovirus type Ⅰ and type Ⅲ.

[0034] (2) Virus inoculation and cultivation: respectively inoculate the duck astrovirus type Ⅰ and type Ⅲ virus species into the cells prepared in step ① according to the final volume of 1:1000, and absorb the virus solution at 37°C for 60 minutes, discard the virus liquid, and 2% newborn bovine serum in DMEM at 37°C, 5% CO 2 Culture until the cells are terminated when lesions appear.

[0035] (3) Virus collection, concentration and purification: freeze-thaw the above-mentioned inoculated diseased cells, collect the supernatant after centrifugation to obtain the virus stock solution; concentra...

Embodiment 3

[0038] Preparation:

[0039] (1) Digest and disperse the BHK-21 cells with trypsin, add 7% newborn bovine serum to the DMEM culture solution in a microcarrier reactor at 37°C and 5% CO 2 After culturing to a single layer of cells, the original culture medium was discarded, and the cells were washed 3 times with serum-free DMEM medium containing 2mM glutamine for inoculation of duck astrovirus type Ⅰ and type Ⅲ.

[0040] (2) Virus inoculation and cultivation: respectively inoculate the duck astrovirus type Ⅰ and type Ⅲ viruses into the cells prepared in step ① according to the final volume of 1:500, and absorb the virus solution at 37°C for 45 minutes, then discard the virus solution in the 1.5% newborn bovine serum in DMEM at 37°C, 5% CO 2 Culture until the cells are terminated when lesions appear.

[0041] (3) Virus collection, concentration and purification: freeze-thaw the above-mentioned inoculated diseased cells, collect the supernatant after centrifugation to obtain th...

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Abstract

The present invention discloses a duck astrovirus vaccine which is an inactivated vaccine obtained by inoculating duck astrovirus type I and type III virus seeds on a continuous cell line BHK-21 cell.The duck astrovirus type I and type III are propagated by the BHK-21 passage cell, and the virus titer is stable to 106.5-107.0TCID50/0.1mL by a Reed-Muench method. The stable production and high titer of antigen are the most important factors in the preparation of the vaccine, the virus titer of duck astrovirus type I and type III proliferation with the BHK-21 cell is more than 10 times that ofa conventional duck embryo method and a primary cell method. If a bioreactor is used for virus proliferation, the virus titer can reach 100 times. The duck astrovirus (type I and III) inactivated vaccine disclosed in the invention has high yield and stable quality, and the immune duck can produce a high-level serum neutralizing antibody, the immune effect is good, and the application prospect is great.

Description

technical field [0001] The invention relates to a duck astrovirus vaccine and a preparation method thereof. Background technique [0002] Astrovirus is a virus that infects mammals and birds. In recent years, the phenomenon of mixed infection with other viruses has become more and more common. Great threat. The family Astroviridae is divided into two genera, Mammalian Astrovirus and Avian Astrovirus, which includes Duck Astrovirus, which is the main cause of duck viral hepatitis (DHV). One of the pathogens, duck astrovirus can cause duck hepatitis, diseased ducklings often have opisthotonos, kidneys are often enlarged, livers are extensively hemorrhaged, and the mortality rate of ducks can reach 50%. Because it can cause high mortality in ducklings, it is an important infectious disease that endangers the duck industry. [0003] At present, there is no duck astrovirus vaccine on the market, and there is no related drug to control the disease caused by the virus. The occur...

Claims

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Application Information

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IPC IPC(8): A61K39/12A61K39/295A61P31/14A61P1/16
CPCA61K39/12A61P1/16A61P31/14A61K2039/5252A61K2039/552A61K2039/70C12N2770/12034
Inventor 张毓金严悌昆敖艳华刘为和谢秉超
Owner 广东渔跃生物技术有限公司
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