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Poliomyelitis inactivated vaccine and production method thereof

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: 2015-01-28
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 production method thereof
  • Poliomyelitis inactivated vaccine and production method thereof
  • Poliomyelitis inactivated vaccine and production method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] Example 1. Experiments on Culture of Poliomyelitis Virus in Basket Reactor under Different Culture Medium and pH Value Conditions

[0037] 1 The experiment of cultivating poliovirus in different medium and different pH value of each medium

[0038] Three different media, M199, DEME and MEM, were used to prepare cell growth medium and virus maintenance solution, and each medium was prepared into four pH gradients, namely pH7.2, pH7.4, pH7.6 and pH7.8. Inoculate Vero cells for culture. After the cells are cultured for 6-8 days and grow into a dense monolayer, wash three times with Earle's solution, and inoculate poliovirus types I, II, and III at the same MOI (MOI=0.01) for culture. , Determination of virus titers and Vero cell residual protein (HCP) content in different culture time.

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

[0040] When type I poliovirus was cultured in three different mediums of M199, DEME and MEM, the viru...

Embodiment 2

[0063] Embodiment 2. Culture experiment of different culture temperature and MOI of cultivating poliovirus in basket reactor

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

[0065] Firstly, the static culture test was carried out in a disposable cell square flask. Referring to the culture temperature of oral poliovirus (33±0.5°C), two different temperatures of 33±0.5°C and 31±0.5°C were selected for the culture of poliovirus. 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 experiment of poliovirus type Ⅰ at different culture temperatures and MOI

[0067] Under the culture condition of 33°C, the virus titer of poliovirus type Ⅰ was higher than that of 31°C, and the level of HCP was basically the same; under the three different MOI culture conditions selected, the virus multiplication peak could...

Embodiment 3

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

[0094] The basket reactor adopts three different dissolved oxygen (DO) setting values ​​of 30%, 50% and 70%, corresponding to the dissolved oxygen (DO) ranges of 20-40%, 40-60% and 60-80%. Three types of poliovirus were cultured, and the change of virus titer under different dissolved oxygen saturation was compared to determine the optimum 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 those when the DO was 50% and 70%; % virus titer has no significant difference, so it is determined that the optimum range of dissolved oxygen is 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%

D...

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

Claims

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Application Information

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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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