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Cat kidney cell line F-81 S adaptive to full suspension cultivation, and applications thereof

A cat kidney cell, F-81S technology, applied in urinary tract/kidney cells, animal cells, vertebrate cells, etc., can solve the problems of limited large-scale application, limited increase in cell density, and inability to maximize growth, etc. Achieve good industrial application prospects, solve the requirements of large-scale industrialization, and achieve the effect of high degree of automation

Active Publication Date: 2017-10-27
郑州爱科生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] With the development of the veterinary vaccine industry, the vaccine preparation technology has also developed from the traditional spinner bottle technology that has been followed for several years to the bioreactor suspension technology. Many patents disclose the use of paper carriers or microcarrier cell suspension culture methods to produce viruses. Vaccine method; the application of carrier-dependent suspension culture technology has improved production automation to a certain extent, and overcomes many shortcomings of spinner bottle technology, but it is still adherent culture in essence, and spinner bottle is still needed to provide seed cells, and expensive Carriers, but also increase the steps of carrier culture and digestion, cell growth still needs a medium with high serum content, and cell growth is limited by many factors and cannot maximize growth, the increase in cell density is limited, especially in industrial scale-up. Congenital defects, which limit the large-scale application of the technology

Method used

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  • Cat kidney cell line F-81 S adaptive to full suspension cultivation, and applications thereof
  • Cat kidney cell line F-81 S adaptive to full suspension cultivation, and applications thereof
  • Cat kidney cell line F-81 S adaptive to full suspension cultivation, and applications thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0027] Embodiment 1: Adapt to the expansion culture of the whole suspension culture F-81S cell line

[0028] The F-81S cells that were preserved in the China Typical Culture Collection Center with the preservation number: CCTCC C201799 were revived and subcultured for full suspension culture, and then inoculated into a bioreactor for suspension culture to obtain F-81S cell suspension culture fluid. Proceed as follows:

[0029] ①Take the F-81S cell line frozen in liquid nitrogen, quickly melt it, add it to a shaker flask filled with nutrient solution, culture it at 36-37°C for 60-72 hours, and subculture and enlarge it according to the ratio of 1:3-1:5;

[0030] ② Dilute the F-81S cell line amplified in step ① to a density of 5-10×105 cells / ml, and inoculate it into a bioreactor. The culture temperature is 36-37°C, the pH value is 6.8-7.2, and the dissolved oxygen is 40%-60%, the F-81S cell suspension growth curve is as follows figure 1 shown;

[0031] ③Determination of cell...

Embodiment 2

[0033] Embodiment 2: The method for cultivating canine parvovirus by using bioreactor suspension culture cells

[0034] Cultivate and expand suspension F-81S cells in shake flasks, press 6.5×10 5 cells / ml density, inoculated into the bioreactor, the cell culture conditions are: the working volume of the reactor is 5 liters, the cell culture temperature is 37 ℃, the pH value is 7.0, the dissolved oxygen is 50%, the culture is 72 hours, and the cell density is 44.7 ×10 5 per ml, replace it with virus culture maintenance medium containing 2.0% newborn bovine serum, and inoculate canine parvovirus according to 5% of the volume of virus culture maintenance medium. The virus culture conditions are: temperature 36°C, pH value 7.0, dissolved Oxygen is 45%, and the virus liquid is harvested after 72 hours of cultivation. The virus titer is judged according to the "Regulations of Veterinary Biological Products of the People's Republic of China", and the harvested virus content is 6.5 T...

Embodiment 3

[0035] Embodiment 3: Utilize the method for cultivating feline panleukopenia virus by suspension culture cell of bioreactor

[0036] The F-81S cell suspension culture method is the same as in Example 2. When cultivating for 72 hours, the cell growth medium is replaced with a virus culture maintenance medium. The virus culture conditions are appropriately adjusted according to the difference of the inoculated virus in the specific examples. With reference to Table 2, all viruses The determination of the titer was carried out according to the routine test method, and the test results are shown in Table 2.

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Abstract

The invention discloses a cat kidney cell line F-81 S adaptive to full suspension cultivation and named as F-81 S. The cat kidney cell line is classified and named as cat kidney cell suspension adapted strain F81S and is preserved at June 29th, 2017, with the preservation number of CCTCC NO:C201799. Compared with the prior art, the cell culture vaccine of the F-81 S adaptive to full suspension cultivation is high in virus automation degree, does not require carrier intervention, can be cultured in a suspended manner in serum-free or low-serum culture medium, solves the large-scale industrial requirements of virus cultivation, develops a large-scale production method capable of meeting GMP production technical requirement, and has good industrial application prospects.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a cat kidney cell line F-81 S suitable for full suspension culture and its application. Background technique [0002] With the rapid growth of China's economy, the pet industry has developed rapidly in recent years, and the number of pets has increased exponentially. The prevention of pet diseases has become a common concern, and the pet vaccine industry is facing unprecedented opportunities for development. At present, there are not many pet vaccines commercialized by domestic animal vaccine companies, and the quality is uneven. They mainly rely on imports, and the products are expensive. Many domestic units are starting to develop products in this field. In addition, my country's special economic animals such as mink, fox, and raccoon dog have the largest stocks in the world. With the continuous expansion of the large-scale breeding of my country's special economic ani...

Claims

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Application Information

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IPC IPC(8): C12N5/071C12N7/00C12R1/91C12R1/93
CPCC12N5/0686C12N7/00C12N2710/16051C12N2750/14351C12N2770/16051
Inventor 李少英徐树兰
Owner 郑州爱科生物科技有限公司
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