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Cultivating and preparing method of I-type dengue virus KMB17 cell adapted strain

A technology of dengue virus and adapted strains, applied in microorganism-based methods, biochemical equipment and methods, viruses/phages, etc., to achieve the effects of good antigenicity, huge economic value and social benefits

Inactive Publication Date: 2014-04-02
INST OF MEDICAL BIOLOGY CHINESE ACAD OF MEDICAL SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Human embryonic lung diploid cell KMB17 has been successfully used in the production of hepatitis A vaccine and polio vaccine. Its safety and cell stability have been widely verified, but it has not been successfully applied to the Chinese strain of dengue virus type I. report

Method used

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  • Cultivating and preparing method of I-type dengue virus KMB17 cell adapted strain
  • Cultivating and preparing method of I-type dengue virus KMB17 cell adapted strain
  • Cultivating and preparing method of I-type dengue virus KMB17 cell adapted strain

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Embodiment 1

[0032] 1) Aedes albopictus cells C6 / 36 were subcultured as follows: Add the following cell growth medium to the cells as usual: MEM medium + 5% volume ratio (W / V) fetal bovine serum + 3.5% volume ratio ( The volume concentration of W / V) is 7% NaHCO3, the volume concentration of 1% volume ratio (W / V) is 3% L-glutamine, the antibiotic of 1% volume ratio (W / V); for 33 + 0.5°C, in an environment with a carbon dioxide volume concentration of 5% (v / v), culture at a constant temperature, change the medium once every 2-3 days, and grow cells into a dense monolayer with good shape after 4-5 days;

[0033] 2) According to the amount of 1ml virus solution / bottle, the number of cells in each bottle is 1~1.7×10 6 , Inoculate type I dengue virus liquid with a virus concentration of 4.0 MOI on the densely growing and transparent Aedes albopictus cells C6 / 36 in step 1), at a temperature of 33 + 0.5°C, CO 2 In an environment with a volume concentration of 5% (v / v), adsorb at a constant temp...

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Abstract

The invention discloses an I-type dengue virus KMB17 cell adapted strain and a preparation method thereof. A popular I-type dengue virus strain can be adapted onto a human embryo lung cell KMB17 for growing to obtain the cell adapted strain which stably goes down to posterity on the KMB17 cell and is strong in virus replication capacity, and the purified adapted strain with high purity is obtained by virtue of a method of purifying bacteriophage plaque by many times. According to a series of identifications on the virus, the I-type dengue virus keeps good antigenicity and basic biological characteristics of the original virus strain. According to the invention, the I-type dengue virus KMB17 cell adapted strain can carry out I-type dengue virus amplification by taking the human embryo lung cell KMB17 as substrate, breaks through limitation of an I-type dengue virus host cell, and lays a foundation for quickening developing a novel idea for researching a dengue virus prophylactic vaccine, and developing I-type dengue virus inactivated and attenuated live vaccines; meanwhile, large-scale production feasibility is achieved.

Description

[0001] technical field [0002] The invention relates to a method for preparing a virus strain, in particular to a method for isolating, screening, breeding, purifying and cultivating an adapted strain of type I dengue virus on KMB17 cells, and belongs to the technical field of virus vaccine manufacture. Background technique [0003] Dengue virus is transmitted by insect vectors such as Aedes aegypti and Aedes albopictus, and is widely prevalent in tropical and subtropical countries and regions. It can cause dengue fever, dengue hemorrhagic fever and dengue hemorrhagic shock syndrome. According to different antigenicity, dengue virus can be divided into four serotypes: I, I, III, and IV. In recent years, due to factors such as global warming and frequent population movements, the incidence of dengue fever in the world has increased by nearly 30 times, and the number of people infected with dengue fever in the world has continued to increase in the past two years. Dengue fev...

Claims

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Application Information

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IPC IPC(8): C12N7/00C12N7/02C12R1/93
Inventor 孙强明赵玉娇陈俊英潘玥黄新伟李多丁晓洁
Owner INST OF MEDICAL BIOLOGY CHINESE ACAD OF MEDICAL SCI
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