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Novel coronavirus vero cell inactivated vaccine virus solution and production method thereof

A coronavirus and production method technology, applied in the field of novel coronavirus Vero cell inactivated vaccine virus liquid, can solve the problems of no novel coronavirus liquid technical report, no effective preventive vaccine for novel coronavirus, etc., and achieve batch-to-batch differences Highly controllable, uniform quality, high cell seeding density

Active Publication Date: 2021-06-18
BEIJING BIOLOGICAL PROD INST CO LTD +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] At present, there is no effective preventive vaccine against the new coronavirus, and no technical reports have been found on the application of basket bioreactors to culture Vero cells to prepare the new coronavirus liquid

Method used

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  • Novel coronavirus vero cell inactivated vaccine virus solution and production method thereof
  • Novel coronavirus vero cell inactivated vaccine virus solution and production method thereof
  • Novel coronavirus vero cell inactivated vaccine virus solution and production method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0049] Embodiment 1: the culture of Vero cell

[0050] Inject a certain volume of cell growth solution into the pre-sterilized 10L basket reactor equipped with a three-dimensional book-type carrier (carrier model: TQ1000w, no-cleaning; provider: Shanghai Chukun, 30-50g / L) (M199 medium containing 8% newborn bovine serum), then inject Vero cells into the tank body, and the cell seeding density is not lower than 1×10 6 cells / ml; then supplement the liquid level with cell growth liquid to the highest scale line of the liquid level and set the parameters of the reactor: the culture speed is 50-150rpm; the dissolved oxygen is not less than 40%; the gas flow rate is 4Gas: 0.1 ; The pH value is controlled between 7.2 and 7.4; the temperature is 36.5±1°C. During the cell culture process, glucose monitoring was used to determine whether the cells grew into a monolayer, and Table 1 was the glucose consumption of Vero cells under different culture time conditions; figure 1 Shows glucose...

Embodiment 2

[0054] Embodiment 2: the cultivation of novel coronavirus

[0055] When the Vero cells were cultured for 6-8 days, the new coronavirus SARS-CoV-2 was inoculated, and the MOI of the three batches of virus inoculation were 0.005, 0.01, and 0.3, respectively. Before virus inoculation, the cell growth liquid in the reactor was discarded, and the cell maintenance liquid (M199 medium without serum) was added to wash and remove bovine serum. Melt the virus seeds in a water bath in the isolator, add the virus seeds into the cell maintenance solution (M199 medium without serum), shake and mix well, then pump the virus-containing cell maintenance solution into the tank. After inoculation, aeration and agitation culture is carried out at 36±1°C, the culture speed is 50-150 rpm, the dissolved oxygen is not less than 40%, and the pH value is controlled between 7.2-7.6 during the culture. Table 2 shows the virus titers of the new coronavirus under different culture time conditions; figur...

Embodiment 3

[0059] Embodiment 3: Harvesting of virus liquid

[0060] From the beginning of virus inoculation, the virus liquid was harvested within 48-96 hours of culture. Turn off the temperature, pH, and DO functions of the basket reactor, open the harvesting pipeline, and use the positive pressure in the tank to collect the virus liquid. The virus titers of the specific virus harvest liquid are shown in Table 3.

[0061] table 3:

[0062] Batches of virus fluid Virus titer (LgCCID 50 / ml)

[0063] From the above experimental results in Table 3, it can be seen that the present invention adopts a basket reactor to cultivate novel coronavirus, with high cell inoculation density, high virus titer, highly controllable batch-to-batch variation, and uniform quality.

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Abstract

The invention provides a novel coronavirus Vero cell inactivated vaccine virus solution and a production method thereof. The method comprises: adding cell growth liquid to a basket bioreactor filled with sheet-like carriers, inoculating Vero cells for culture, and then washing with cell maintenance liquid after growing into a dense monolayer; injecting cells to maintain The MOI of the virus inoculation is 0.005-0.4, the virus culture temperature is 36±1°C, the dissolved oxygen is ≥40%, and the pH value of the virus culture stage is controlled to be 7.2-7.6; after the virus inoculation, cultivate for 48 ~96h, the virus fluid was harvested. In the present invention, the novel coronavirus Vero cell inactivated vaccine virus liquid is successfully cultivated by using a basket reactor. The present invention has high cell inoculation density, high virus titer, highly controllable batch-to-batch difference, and uniform quality; it is a new type of coronavirus inactivated The large-scale production of vaccines has laid a solid foundation.

Description

technical field [0001] The invention relates to a novel coronavirus Vero cell inactivated vaccine virus liquid and a production method thereof, specifically a method for producing a novel coronavirus inactivated vaccine virus liquid using a basket bioreactor using Vero cells as a matrix, and utilizing The method produces the novel coronavirus Vero cell inactivated vaccine virus liquid. Background technique [0002] A new type of coronavirus appeared in 2019, which was confirmed by the World Health Organization (WHO) and named as SARS-CoV-2, and the pneumonia caused by this pathogenic infection is called new type of coronavirus pneumonia. At present, there is no effective drug for the treatment of new coronary pneumonia, and the development of a vaccine for new coronary pneumonia is imminent. [0003] SARS-CoV-2 is a new type of coronavirus with an envelope, and the particles are round or oval, often pleomorphic, with a diameter of 60-140nm. The new coronavirus is another c...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N7/00A61K39/215A61P31/14A61P11/00C12R1/93
CPCA61K39/12A61K2039/5252A61P11/00A61P31/14C12N7/00C12N2770/20034C12N2770/20051
Inventor 王辉赵玉秀付慧李婉莉李爱灵张越于守智张征丽郭彦岑杨晓明
Owner BEIJING BIOLOGICAL PROD INST CO LTD
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