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CV-A10 virus species and inactivated vaccine thereof for human

A CV-A10, inactivated vaccine technology, applied in the direction of positive-sense single-stranded RNA viruses, viruses, antiviral agents, etc., can solve problems such as difficulty in control, and achieve the effect of increasing production efficiency

Pending Publication Date: 2019-03-29
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

Although the EV71 vaccine has been successfully marketed and used for the prevention and treatment of hand, foot and mouth disease, HFKM is caused by a variety of enteroviruses. In order to solve the problem that EV71 vaccine is difficult to control other enteroviruses such as CV-A10 and other enteroviruses that cause HFMD and possible EV -A71 vaccine public acceptance and questioning issues, it is necessary to develop HFMD vaccine of CV-A10

Method used

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  • CV-A10 virus species and inactivated vaccine thereof for human
  • CV-A10 virus species and inactivated vaccine thereof for human
  • CV-A10 virus species and inactivated vaccine thereof for human

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0044] Embodiment one: the isolation, adaptation and purification method of the virus strain involved in the present invention

[0045] (1) Obtain virus samples

[0046] Collect excreta samples from hand, foot and mouth disease patients identified as CV-A10 infection by the Pediatric Department of the hospital, take 1g of feces and resuspend in 5mL of normal saline, centrifuge at 3000g for 30 minutes, take the supernatant, and use a 0.45μm filter (purchased from Millipore Company) Filter and store at -20°C for later use.

[0047] (2) Adaptation of virus on RD cells

[0048] Select the RD cells that have grown into a dense monolayer, discard the old culture medium, wash the cell surface with an appropriate amount of PBS solution, and add an appropriate amount of 0.125% trypsin to digest the cells. Inoculate 2ml of the culture medium into each well of a 24-well plate, inoculate 200μl of the filtered sample on the cell surface, incubate at 37°C for 3 to 4 days, observe the cyto...

Embodiment 2

[0055] Embodiment 2: Cultivation of CV-A10 strain and vaccine preparation method

[0056] (1) The virus was cultured in KMB17 cells

[0057] Select the P28 generation KMB17 cells that have grown into a dense monolayer, discard the old culture medium, wash the cell surface with an appropriate amount of PBS solution, add an appropriate amount of 0.125% trypsin to digest the cells, and gently pat the cell bottle, the cells slide down from the bottle wall like quicksand, Add cell culture medium, press 2×10 5 Inoculate the small square bottle with cells / bottle, inoculate at 37°C for 3 to 4 days, inoculate 200 μl of CV-A10 virus on the cell surface, adsorb at 37°C for 1 hour, then add MEM cell maintenance solution without calf serum, incubate at 37°C and Observe the cell lesions and the virus CPE reaches 90%. Harvest the virus and freeze it for later use. attached Figures 1a-1d For the CV-A10 strain to produce pathological changes in KMB17 cells (cultured for 4 days), cytopathic...

Embodiment 3

[0064] Embodiment three: the identification method of virus seed

[0065] (1) Sterility inspection and mycoplasma detection experiment

[0066] Take 4 tubes of thioglycolate liquid medium and 2 tubes of tryptone soy liquid medium, and inoculate each tube with 0.5 ml of virus. After inoculation, two thioglycolate fluid mediums were cultured at 30-35°C, two thioglycolate fluid mediums and two tryptone soytone liquid mediums were cultured at 20-25°C; The method was used as a negative control; the results were judged after 14 days of culture. Take 4 tubes of semi-solid medium and broth medium for mycoplasma inspection, boil and melt the semi-solid medium, cool to 56°C, add energy-killed calf serum mixed in a ratio of 4:1 to each of the two media and 2.5ml of double-antibody mixture, shake quickly, and then add 0.5ml of the sample to be tested to each culture medium for primary culture, and cultivate and observe at 35°C; on the 7th day, take out 2 culture mediums for each of the ...

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Abstract

The invention discloses a CV-A10 virus species and an inactivated vaccine thereof for human. The classification name of the species is coxsackie virus A group 10-type with the preservation number being CGMCC No.16218; and the prepared CV-A10 inactivated vaccine for human consists of 100 [mu]g / ml of CV-A10 inactivated purified antigens and 1mg / ml of aluminium hydroxide. Results indicate that the vaccine products have favorable immunogen and safety.

Description

technical field [0001] The invention relates to a CV-A10 virus seed and an inactivated vaccine for human use. Background technique [0002] Hand foot mouth disease (hand foot mouth disease, HFMD) in the world, especially in the Asia-Pacific region, the epidemic scale and threat impact are increasing. So far, the epidemiological statistics in my country show that since the disease was listed as a Class C infectious disease in 2008, it has become the first in the number of cases of Class C infectious diseases in 2009, and the overall case fatality rate is about 0.05%. The prevention and control of this infectious disease, especially among children, has become an important public health issue. [0003] In recent years, CV-A10 (Coxsackievirus A group A type 10) infection has an increasing tendency to cause HFMD outbreaks or epidemics, and some areas have become the main pathogens of HFMD other than EV71, CV-A16 and CV-A6. Several other enteroviruses, including EV71, CV-A16, an...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N7/00A61K39/125A61P31/14
CPCA61K39/12A61P31/14C12N7/00C12N2770/32021C12N2770/32334A61K2039/5252
Inventor 马绍辉张捷刘红波张名张海浩赵义林杨昭庆黄小琴孙浩
Owner INST OF MEDICAL BIOLOGY CHINESE ACAD OF MEDICAL SCI
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