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Poliomyelitis inactivated vaccine and its production method

A technology for polio and inactivated vaccine, which is applied in the field of inactivated polio vaccine and its production, can solve the problems of low virus titer in harvested liquid and no disclosure of culture conditions, and achieves high cell inoculation density, uniform quality, Highly controllable effect of batch-to-batch variation

Active Publication Date: 2017-09-19
BEIJING BIOLOGICAL PROD INST CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

CN102406928A discloses a production method of inactivated human diploid cell polio vaccine, wherein multi-stage microcarrier bioreactors are used to amplify and subculture human diploid cells, and the cell growth of each stage bioreactor reaches 10 6 When the concentration is above 1 / ml, digest the cells to prepare cell suspension, enter the next-level bioreactor for amplification and culture according to the amplification ratio of 1:10-1:20 until the scale required for production is reached, and then inoculate poliovirus , Proliferation culture, harvesting virus culture fluid and performing purification and inactivation, but the virus titer of the harvest fluid produced by this technology is low, and the specific culture conditions are not disclosed
[0006] At present, there are no technical reports on the application of basket bioreactors to efficiently produce high-quality inactivated polio vaccines using Vero cells as a substrate

Method used

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  • Poliomyelitis inactivated vaccine and its production method
  • Poliomyelitis inactivated vaccine and its production method
  • Poliomyelitis inactivated vaccine and its production method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] Example 1. Experiment of culturing poliovirus in basket reactor under different culture medium and pH value conditions

[0037] 1 Experiment of culturing poliovirus under different media and different pH values ​​of each media

[0038] M199, DEME and MEM were used to prepare cell growth solution and virus maintenance solution. Each medium was prepared into 4 pH gradients, pH 7.2, pH 7.4, pH 7.6 and pH 7.8, respectively. Vero cells were inoculated for culturing. After the cells were cultured for 6 to 8 days and grew into a dense monolayer, they were washed three times with Earle's solution, and were inoculated with I, II, and III polioviruses at the same MOI (MOI=0.01) for culture. , the virus titer and the residual protein (HCP) content of Vero cells were determined at different times of culture.

[0039] 1.1 Culture test of type I poliovirus under different medium and pH conditions

[0040] When type I poliovirus was cultured in M199, DEME and MEM, the viral titers of ...

Embodiment 2

[0063] Embodiment 2. Cultivation test of different culture temperature and MOI of poliovirus in basket reactor

[0064] 1. Preliminary screening of poliovirus culture temperature and MOI conditions in disposable cell flasks

[0065] First, static culture experiments were carried out in disposable cell square flasks. Referring to the culture temperature of oral poliovirus (33±0.5°C), two different temperatures, 33±0.5°C and 31±0.5°C, were selected for poliovirus culture. Three different MOIs, namely 0.01, 0.1 and 0.3, were used to detect the virus titer and HCP content at different temperatures and different MOIs.

[0066] 1.1 Culture test of type I poliovirus at different culture temperature and MOI

[0067] Under the culture condition of 33℃, the virus titer of type I poliovirus was better than that of 31℃, and there was basically no difference in the level of HCP; under the three different MOI culture conditions selected, the virus reproduction peak could be reached for 60...

Embodiment 3

[0093] Example 3. Determination of Dissolved Oxygen for Poliovirus Culture in Basket Reactor

[0094] The basket reactors used three different dissolved oxygen (DO) settings of 30%, 50% and 70%, which corresponded to dissolved oxygen (DO) ranges of 20-40%, 40-60% and 60-80%. Three types of poliovirus were cultured, and the changes of virus titer under different dissolved oxygen saturation were compared to determine the appropriate DO for culturing poliovirus in the basket reactor. For other culture conditions, refer to Example 2.

[0095] The test results showed that when the DO was 30%, the virus titers of the three types were far lower than the virus titers when the DO was 50% and 70%. There was no significant difference in % virus titer, so the optimal dissolved oxygen range was determined to be 40-80%. The results are shown in Table 11 to Table 13.

[0096] Table 11 Virus titers of type I poliovirus under different dissolved oxygen

[0097] time

DO = 30% ...

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Abstract

The invention provides a poliomyelitis inactivated vaccine and a production method thereof. The poliomyelitis inactivated vaccine is produced by applying a basket-type bioreactor and adopting Vero cells as a matrix. The production method comprises the following specific steps of adding an M199 culture medium into the basket type reactor, inoculating the Vero cells for culture, and after a compact monolayer is grown, washing and changing with Earle's balance salt solution; after the washing and the changing are finished, injecting M199 culture medium solution, inoculating poliomyelitis virus with virus inoculation MOI being 0.01-0.3, further culturing with the temperature being 33+ / -0.5 DEG C, dissolving oxygen by 40-80%, controlling a pH value of a system at the virus culture stage to 7.4-7.6; and after inoculation, culturing for 42-72 hours, thereby obtaining virus solution. The poliomyelitis inactivated vaccine and the production method provided by the invention have the advantages that the proper culture medium is selected and the conditions of proper culture temperature, pH value, inoculating proportion and dissolved oxygen and the like are controlled, so that the virus titer is high, the antigen content is high, the height for differences between batches can be controlled and the mass is uniform.

Description

technical field [0001] The invention relates to an inactivated polio vaccine and a production method thereof, in particular to a method for producing an inactivated polio vaccine by using a basket bioreactor with Vero cells as a matrix, and a spinal cord produced by the method. Inactivated poliomyelitis vaccine. Background technique [0002] Polio (referred to as polio) is an acute intestinal infectious disease caused by poliovirus infection, mainly limb paralysis. The main susceptible population for poliomyelitis is children. At present, there is still no effective method to treat polio. The most important thing to control and eliminate polio is an effective polio vaccine. [0003] There are two types of polio vaccines that have been used at home and abroad: oral live attenuated polio vaccine (Oralpoliovirus vaccine, OPV) and inactivated poliovirus vaccine (IPV). Among them, IPV does not cause circulating iVDPV infection caused by Vaccine Associated Paralytic Poliomyelit...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N7/00A61K39/13A61P31/14C12R1/93
CPCY02A50/30
Inventor 王辉赵玉秀李爱灵梁宏阳刘小娟赵硕董圆曾令冰
Owner BEIJING BIOLOGICAL PROD INST CO LTD
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