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Rapid adaptation method of rabies vaccine virus strains for human body on diploid cells of human body

A technology of human diploid cells and diploid cells, applied in the field of vaccines, can solve the problems of low titer and long time consumption, and achieve the effects of saving manpower and material resources, increasing virus yield and shortening adaptation time.

Active Publication Date: 2013-12-25
云南沃森生物技术股份有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] All of the above methods are carried out with poisonous passage and need to gradually increase the number of fresh cells. It takes more than 40 passages to adapt, which takes too long and the titer is low

Method used

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  • Rapid adaptation method of rabies vaccine virus strains for human body on diploid cells of human body
  • Rapid adaptation method of rabies vaccine virus strains for human body on diploid cells of human body
  • Rapid adaptation method of rabies vaccine virus strains for human body on diploid cells of human body

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0040] Example 1 Rejuvenation of viruses on susceptible animals

[0041] Take the strains CTN strain-1V5, PM strain and aG strain produced by the rabies vaccine respectively and make 10000-fold dilution, and inoculate 9-11g Kunming mice, 0.03ml / mouse, to the rat brain, and feed for 4-5 days. Choose the ones with typical rabies symptoms The mouse brain was aseptically taken from the mouse. Prepared to 10 -1 Mouse brain suspension, take 10 -1 The virus suspension was diluted to 10 with 2% calf serum phosphate buffer -2 Virus suspension, repeat the above steps for passage, and use after passage for 3 generations.

Embodiment 2

[0042] Example 2 Adaptation of CTN-1V5 virus on Walvax-2 cells and MRC-5 cells

[0043] 2.1 Resuscitate Walvax-2 cells and MRC-5 cells by conventional methods. Use MEM containing 10% calf serum to pass through to 25-30 generations. After growing into a monolayer, they are digested and dispersed into single cells. After adding complete culture medium, they are centrifuged and Remove the supernatant, and suspend the cell pellet in a serum-free culture medium to prepare a human embryonic lung diploid cell suspension dispersed into single cells;

[0044] 2.2 Inoculate the fresh rabies virus CTN-1V5 virus into a suspension of human embryo lung diploid cells dispersed into single cells at 0.0001-1 MOI, adsorb at 37°C for 90 minutes, and add cell culture medium to a suitable volume of 8 ml Incubate at 30-40°C and form a dense monolayer in 2-5 days. Discard the cell culture medium and replace it with MEM medium containing 5% calf serum and 1% sucrose solution for 5 days to harvest the supe...

Embodiment 3

[0051] Example 3 Adaptation of aG strain on Walvax-2 and MRC-5

[0052] Using the adaptation method of Example 2, the aG strain was adapted to the Walvax-2 and MRC-5 cells, and the rat brain method was used for titer detection.

[0053]

[0054] Example 3 Adaptation of CTN-1V5 virus strain on KMB-17 and adaptation of PM strain on WI-38

[0055] Using the adaptation method of Example 2, the CTN-1V5M virus strain was adapted to KMB-17, the PM strain was adapted to WI-38, and the rat brain method was used for titer detection.

[0056]

[0057]

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Abstract

The invention discloses a rapid adaptation method of rabies vaccine virus strains for the human body on diploid cells of the human body. Fresh rabies viruses are inoculated on the diploid cells of the human body, poisoned passage is carried out, only 4-5 cycles are required, passage is carried out for 12-15 passages, the virus titer rises above 7.0logLD50 / ml of the P15 from 3.0logLD50 / ml of the P1, the adaptation cycle of the viruses is shortened by at least 50%, on the premise that the virus immunogenicity is not affected, the virus yield is rapidly improved, and manpower and material resources are saved.

Description

[0001] Field of invention: [0002] The present invention belongs to the technical field of vaccines. Specifically, the present invention relates to a method for rapid adaptation of human rabies vaccine strain on human diploid cells. Background technique [0003] Rabies is a zoonotic natural epidemic disease caused by rabies virus infection. It is an infectious disease with the highest number of deaths and case fatality rates in China's legal reports. According to the 2003 World Rabies Survey Report, the number of cases in China is second only to India, ranking second in the world. The Ministry of Health of China has listed rabies as a Class 2 infectious disease for management, but the rabies epidemic in China has been on the rise in recent years. The epidemic situation is very severe. [0004] The rabies vaccines currently mass-produced in the world include diploid cell rabies vaccine, Vero cell inactivated vaccine, hamster kidney cell, chicken embryo, duck embryo vaccine. Experim...

Claims

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Application Information

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IPC IPC(8): C12N7/00
Inventor 杨喆马波何丽芳王丽丽李伟陈敏王馨尹孝兵
Owner 云南沃森生物技术股份有限公司
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