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Goat inactivated vaccine and preparation method thereof

A technology for inactivated vaccines and goat pox, applied in biochemical equipment and methods, methods based on microorganisms, pharmaceutical formulations, etc., can solve problems such as long production cycle, low virus titer content of virus liquid, and complicated cell culture process, etc. To achieve the effect of avoiding seasonal restrictions, avoiding potential risks, and good immune protection

Inactive Publication Date: 2015-10-07
CHINA ANIMAL HUSBANDRY IND
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  • Abstract
  • Description
  • Claims
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Problems solved by technology

However, the disadvantage of the above-mentioned goat pox attenuated live vaccine is that the cost of using sheep testicular cells to culture the virus is relatively high, limited by the season, and only in spring can a large number of sheep testes be collected, the cell culture process is complicated, and the main reason is that the use of homologous cells The vaccine made by breeding the virus has the phenomenon of recessive virus transmission and virulence reversion after immunizing the sheep body, which makes the current live vaccine use process unsafe
In order to avoid the phenomenon of recessive virus transmission and virulence reversion brought about by culturing goat pox virus with homologous tissue cells, those skilled in the art have been devoting themselves to research on the use of BHK-21 cells to culture goat pox virus, but the results are not ideal. The virus titer in the harvested virus solution is low, the BHK-21 cells have many generations of domestication, and the BHK-21 cells inoculated with goat pox virus need to be passaged many times, resulting in a long production cycle and unsatisfactory virus harvesting effect

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  • Goat inactivated vaccine and preparation method thereof

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Experimental program
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Effect test

Embodiment 1

[0023] The preparation of embodiment 1 goat pox inactivated vaccine

[0024] 1. Preparation of spinner bottle cell virus:

[0025] Take a single layer of well-grown BHK-21 cells, discard the nutrient solution, digest with trypsin, add milk liquid containing 2% calf serum, and inoculate the goat pox virus seed used for production on the BHK-21 digested with trypsin. 21 In the cell suspension, make up the cell maintenance solution, culture at 37°C, and observe the cell changes every day. Viral cultures were harvested after approximately 20 hours when cytopathy reached 75%.

[0026] 2. Suspension cell virus preparation:

[0027] Take 1 branch of working seed cells, resuscitate and adhere to the wall for 2 passages, grow to a monolayer and then culture in suspension for 3 passages in shake flasks, when the cell concentration reaches 2×10 6 / ml or more were transferred to 5L or 100L bioreactors for cell expansion and culture until the cell concentration reached 2×10 6 / ml or mo...

Embodiment 2

[0034] Example 2 Efficacy Test of Goat Pox Inactivated Vaccine

[0035] Screening of goat pox antibody-negative goats: through trial and error, a goat pox cell neutralization test was established: the goat serum to be tested was diluted 1:4 and mixed with 200 TCID 50 The goat pox venom was neutralized at 37°C for 1 hour, and inserted into well-growing bovine testicular cells. At the same time, a virus control, a negative control, and a goat pox positive serum control were set up to observe cell lesions. The virus control showed typical lesions, and the negative control and goat pox positive The serum control had no lesions, and the test was established. If lesions appear in the neutralized sample of the goat serum to be tested, the serum goat pox antibody is negative. Goat pox antibody was tested on the goats used in the experiment, and goats negative for goat pox antibody were successfully screened out.

[0036] According to Announcement No. 442 of the Ministry of Agricultu...

Embodiment 3

[0037] Example 3 Safety Test of Goat Pox Inactivated Vaccine

[0038] According to Ministry of Agriculture Announcement No. 442 "Veterinary Biological Products Registration Classification and Registration Data Requirements", the safety inspection of the 3 batches of vaccines prepared in Example 1 was carried out as follows:

[0039] Each batch of goat pox antibody-negative goats was vaccinated with 2 goats, 3ml / goat, injected subcutaneously in the ventral part, and the sheep were vaccinated, causing slight redness and swelling locally, which subsided after 3-5 days, observed for 15 days, and no secondary pox swelling appeared .

[0040] Healthy guinea pigs with a body weight of 350-450 g were subcutaneously injected with 2 ml each and healthy mice with a body weight of 18-22 g were each subcutaneously injected with 0.5 ml, and observed for 7 days.

[0041] Each batch of inactivated antigen injection goat pox antibody negative 3 goats, ventral intradermal injection, 1ml / goat, ...

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Abstract

The invention provides a goat inactivated vaccine and a preparation method thereof. The preparation method comprises the following steps: goat pox AV41 is inoculated with BHK-21 passage cells, as a result, CPE is generated, continuous inoculation is carried out to harvest antigens, BHK-21 cell venom is inactivated, and the goat pox BHK-21 inactivated vaccine, of which both the efficacy and safety are inspected to be qualified is finally prepared through a preparation technology for exploring inactivated vaccine. According to the invention, goat testicle cells are replaced with the BHK-21 cells, so that the potential risk that the vaccine carries pathogene when homologous tissue is used for preparing the vaccine is avoided completely, and meanwhile, seasonal limitation that a large number of goat testicles can be collected in spring only is avoided; since the culture cost of the BHK-21 cell is lower that of the beef and goat testicle cell, the goat pox BHK-21 inactivated vaccine provided by the invention is safe and reliable, can be used for preventing and treating goat pox, is low in preparation cost and has better social benefits and economic benefits.

Description

technical field [0001] The invention relates to the field of vaccine preparation, in particular to a goat pox BHK-21 cell inactivated vaccine and a preparation method thereof. Background technique [0002] Goat pox, also known as "sheep smallpox", is an acute, febrile, contact infectious disease caused by goat pox virus. The disease can occur in any season, but it is more common in spring and autumn. The morbidity and mortality of the disease are very high, which seriously restricts the development of goat breeding industry. At present, the most effective way to control this disease is regular vaccination. China Animal Husbandry Group Lanzhou Biopharmaceutical Factory first developed goat pox tissue vaccine, and later successfully developed goat pox attenuated cell vaccine, which won the Development Award of the Ministry of Agriculture. Tens of millions of doses of goat pox attenuated live vaccine are produced every year for immune prevention. The existing goat pox attenu...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K39/275A61P31/20C12N7/00C12R1/93
Inventor 吕红沈芳郑明举马爱荣孔繁梅张爱民
Owner CHINA ANIMAL HUSBANDRY IND
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