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Preparation method of newcastle disease, avian influenza virus and avian adenovirus trivalent inactivated vaccine

A technology of chicken Newcastle disease virus and avian adenovirus, applied in biochemical equipment and methods, vaccines, viruses, etc., can solve the problems of difficult to achieve high-efficiency transfection, unfavorable experimental operation, cumbersome preparation, etc., and achieve complete immune protection and antibody production Fast, long-lasting effect

Inactive Publication Date: 2017-05-31
广州博恒生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the avian adenovirus is cultured with primary chicken hepatocytes, which is cumbersome to prepare, easily polluted, and difficult to achieve high-efficiency transfection, which is not conducive to experimental operations.

Method used

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  • Preparation method of newcastle disease, avian influenza virus and avian adenovirus trivalent inactivated vaccine
  • Preparation method of newcastle disease, avian influenza virus and avian adenovirus trivalent inactivated vaccine
  • Preparation method of newcastle disease, avian influenza virus and avian adenovirus trivalent inactivated vaccine

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] The preparation of embodiment 1 liposome complex:

[0042] (1) Weigh 0.2g of phosphatidylcholine, 0.08g of cholesterol, and 0.04g of distearoylphosphatidylethanolamine, mix them, dissolve in 50mL of ethanol, and sonicate for 5min;

[0043] (2) Rotate evaporation at 28°C and 0.1MPa to remove ethanol, let stand for 30min, add 100mL phosphate buffer solution containing 0.5% (m / v) catalase at pH 7.4, and place in a water bath at 35°C Medium shaking hydration reaction for about 1 hour to obtain liposome suspension;

[0044] (3) Adjust the particle size of the liposome suspension to 150 nm using a syringe filter to obtain liposomes containing catalase inside.

Embodiment 2

[0045] Example 2 Preparation of poultry adenovirus type 4 seedling virus solution and determination of virus content

[0046] (1) Preparation of poultry adenovirus type 4 seedling virus liquid:

[0047] ① Cell preparation: Digest and disperse LMH cells with trypsin-0.02% EDTA with a mass volume ratio of 0.025%, add 10% (v / v) newborn bovine serum, 1.0% (v / v) double antibody and 2.0% (m / v) DMEM culture solution of hydroxyethylpiperazineethanesulfonic acid, at 37°C, 5% CO 2 After culturing to a monolayer of cells, wash the cells three times with serum-free DMEM medium for avian adenovirus type 4 virus inoculation;

[0048] ②Inoculation and cultivation of avian adenovirus type 4: inoculate the cells prepared in step ① with the virus seed of avian adenovirus type 4 at a final volume of 1:200, absorb the virus solution at 37°C for 40 minutes, and add 1.0% (v / v) newborn bovine serum, 0.4% (m / v) glutamine, 1.0% (v / v) double antibody, 2.5% (v / v) liposome complex, 0.15% (m / v) Lipoic ...

Embodiment 3

[0060] Example 3 Preparation of virus solution for chicken Newcastle disease vaccine production and avian influenza H9 subtype vaccine production and virus content determination

[0061] (1) Preparation of Newcastle Disease Vaccination Virus Liquid:

[0062] Dilute chicken Newcastle disease virus seed to 10 with sterilized PBS -4 or 10 -5 , inoculate 10-11-day-old chicken embryos, inoculate 0.2ml into the allantoic cavity of each embryo, and continue to incubate at 36-37°C. It is not necessary to turn the embryos, and irradiate the embryos once a day, and incubate them for 96 hours. Take them out, irradiate the embryos, and discard them. Remove the dead embryos, put the air chamber of the living embryos upright, and cool at 2-8°C for 12-24 hours, take out the cooled chicken embryos, harvest the chicken embryo allantoic fluid, absorb the chicken embryo allantoic fluid and place it in a sterilized container, Take a sample to measure the agglutination value of red blood cells. ...

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Abstract

The invention relates to a preparation method of a newcastle disease, avian influenza virus and avian adenovirus trivalent inactivated vaccine. The method comprises the following steps: (1) inoculating an avian adenovirus type 4 virus into an LMH cell and obtaining a vaccine-made virus liquid through the steps of culture, freeze thawing, centrifugation and concentration; (2) inoculating a newcastle disease virus La Sota strain and an H9 subtype avian influenza virus into a chick embryo allantoic cavity separately, culturing to obtain a chick embryo allantoic liquid and carrying out centrifugation and concentration to obtain a newcastle disease and H9 subtype avian influenza virus vaccine-made virus liquid; and (3) inactivating virus liquids of the three viruses, and carrying out isopyknic mixing and emulsification to obtain an emulsion-type inactivated vaccine. The trivalent inactivated vaccine provided by the invention has a complete immune protecting effect on the newcastle disease virus, the H9 subtype avian influenza virus and the avian adenovirus type 4 virus, and is good in security, stable, effective and long in protection period.

Description

technical field [0001] The invention belongs to the technical field of veterinary biological products, and in particular relates to a preparation method of a triple inactivated vaccine of Newcastle disease, avian influenza virus and avian adenovirus. Background technique [0002] Newcastle disease (ND), commonly known as chicken plague, is a highly contagious, acute, and severe infectious disease of chickens caused by Newcastle disease virus. It has a high morbidity and fatality rate. Industry is devastated. [0003] H9N2 subtype avian influenza belongs to low pathogenicity avian influenza with high morbidity but low mortality. The main manifestations are only mild respiratory symptoms, reduced feed intake, egg production rate can drop from 90% to less than 20%, or even go out of production, and about 30% of commercial broilers can die. It is very easy to be mixed with E. coli infection, serious It affects the production performance of poultry and brings serious economic l...

Claims

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Application Information

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IPC IPC(8): A61K39/17A61K39/145A61K39/235A61K9/107A61P31/20A61P31/16A61P31/14C12N7/00
CPCA61K39/12A61K9/107A61K2039/5252A61K2039/70C12N7/00C12N2710/10234C12N2710/10251C12N2760/16134C12N2760/18134
Inventor 张毓金谢秉超严悌昆
Owner 广州博恒生物科技有限公司
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