MDBK acclimation suspension method and two-stage virus production process

Abstract

The invention relates to an MDBK cell acclimation suspension method and a two-stage virus production process. According to the MDBK cell acclimation suspension method, adherent MDBK cells are acclimated to adapt to serum-free suspension culture and used as host cells for producing viruses which can have cytopathic effect or sensitivity on suspension MDBK cells by two-stage culture virus inoculation; when the cells grow to 2.0-20.0*10<6> cells / mL, diluting with a production culture medium by 1-5 times is performed to reach a density range of 1.0-10.0*10<6> cells / mL, and then virus inoculation is carried out or virus inoculation is performed before dilution. The suspension MDBK cells are adaptive to suspension growth in the serum-free chemical-defined culture medium, and the two-stage virusproduction process is simple and easy in amplification; liquid changing is avoided, and high culture medium utilization efficiency is achieved; production and growth culture media can be identical ornot and can be mixed proportionally to realize cell secondary growth and viral expression promotion.

Description

technical field [0001] The invention relates to the field of MDBK cell acclimation suspension method and suspension MDBK cell-related vaccine production. The MDBK acclimation suspension method and the second-stage virus production process of the present invention can exist simultaneously or independently in the technical scheme. Specifically, it relates to a process technology for cultivating suspended MDBK cells in a bioreactor above 30L and cultivating the suspended MDBK cells in a second-stage culture method to produce vaccines. Background technique [0002] Mammalian cell large-scale culture technology is one of the downstream common technologies of biopharmaceutical companies, and it plays an extremely important role in this industry. The key to this technology is to achieve large-scale culture of mammalian cells in suspension serum-free, thereby increasing production capacity, reducing costs, and facilitating scale-up. The usual practice in culture of adherent cells i...

AI Technical Summary

Problems solved by technology

[0004] The defects of the existing MDBK cell acclimatization and suspension are: 1. Some cannot achieve complete serum-free culture; 2. Generally, a two-step method is adopted, that is, first reduce to serum-free and then suspend culture, because the medium used for adherent culture Serum is usually added to the basal medium. The nutrient content of the basal medium is weak, and the surface area / volume ratio of the adherent culture during the domestication process is usually small, which limits the intake of nutrients. The entire domestication process takes a long time; 3. After genetic modification, domestication, That is, through genetic modification, insert gene fragments that are not conducive to adherence into the adherent cell line, change the original adherent culture method, and make the cells grow in suspension, and then through continuous optimization and change of the medium, the cells can be serum-free and high-density. Growth, this method is easy to implement, but the insertion of foreign gene fragments affects the quality of the product

Up to now, there are no related reports on the second-stage culture method of suspension MDBK cells

Images