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A kind of dual combination hand-foot-mouth disease inactivated vaccine and preparation method thereof

A technology of virus strains and microbial strains, applied in the biological field, can solve the problems of shortening fever and skin lesion healing time, difficulty in finding and controlling the source of infection, and difficulty in effectively blocking, etc., achieving good application prospects and extensive neutralization and protection , the effect of low production cost

Inactive Publication Date: 2018-05-04
JILIN UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Hand, foot and mouth disease caused by EV71 and CoxA16 has the characteristics of recessive infection and mild cases, the source of infection is difficult to find and control, children are generally susceptible, and there are many transmission routes, which are difficult to effectively block. High-efficiency and broad-spectrum antiviral drugs such as acyclovir, valacyclovir, and famciclovir are commonly used to shorten fever and skin lesion healing time, and relieve oral herpes pain. So far, there is no specific drug for the prevention of this disease

Method used

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  • A kind of dual combination hand-foot-mouth disease inactivated vaccine and preparation method thereof

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preparation example Construction

[0030] A preparation method of dual hand, foot and mouth disease inactivated vaccine, comprising the following steps:

[0031] Step 1: The human intestinal 71 virus strain CC-09 with the deposit number CGMCC No.7753 and the Coxsackie A16 virus strain CC-16 with the deposit number CGMCC No.7752 were respectively inoculated on Vero cells expanded and subcultured Or cultured in human embryonic lung diploid cells;

[0032] Step 2: Harvest the two kinds of virus culture fluids separately, concentrate, inactivate, and purify after removing impurities to obtain two kinds of virus stock solutions;

[0033] Step 3: Sterilize the two virus stock solutions prepared in step 2, and mix them to obtain the dual-combination hand, foot and mouth disease inactivated vaccine.

[0034] The preparation method of the dual-combination hand, foot and mouth disease inactivated vaccine of the present invention firstly inoculates the two virus strains into expanded and subcultured Vero cells or human emb...

Embodiment 1

[0055] In order to further understand the present invention, the present invention will be described in detail below in conjunction with examples. Embodiment 1: the screening of virus strain and the mensuration of the virulence of virus strain

[0056] 1. Virus strain screening:

[0057] A total of 15 strains of EV71-positive virus and 20 strains of CA16-positive virus were isolated from the throat swabs of hand, foot and mouth patients. They were respectively adapted to Vero culture, and their biological characteristics were analyzed and studied one by one. Large and typical plaques were selected. 2-3 virus strains with high infectious titer and wide range of cross-protection are used as candidate strains for the first round of vaccines.

[0058] Plaque cloning and purification of the first round of vaccine candidate strains were carried out, multiple clones were selected for full sequence determination, the original seed batch was selected to establish the original seed bat...

Embodiment 2

[0065] Embodiment 2: the preparation of the dual hand-foot-mouth inactivated vaccine of the present invention

[0066] (1) Vaccine preparation: take the African green monkey kidney cells (Vero) cultured in a spinner bottle covered with a monolayer, digest with 0.25% trypsin (containing EDTA), inoculate a bioreactor according to a certain number of cells, and cultivate for 3-5 days Take working seeds of EV71 or CA16 strains and batch virus seeds, inoculate Vero cells in the bioreactor after appropriate dilution, culture at 35°C±0.5°C for 72-120 hours, harvest the virus culture medium, freeze and thaw in a -20°C freezer, and 0.1 μm ~ 0.5 μm pore size filter to remove cell debris, 300KD molecular weight cut-off ultrafiltration membrane concentrated 50 times, under sterile conditions, add formaldehyde with a final concentration of 1:4000 to inactivate at 37°C for 10 days, and the inactivated virus solution, equilibrated with 0.02mol / L PB buffer (pH 5.0-7.0), purified by anion exch...

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Abstract

The invention relates to the field of biotechnology, and discloses two hand, foot and mouth disease virus strains and a dual hand, foot and mouth disease inactivated vaccine prepared from the two virus strains. The human intestinal 71 type virus strain CC-09 with the preservation number CGMCC No.7753 and the Coxsackie A16 virus strain CC-16 virus with the preservation number CGMCC No.7752 were respectively inoculated in Vero cells that were expanded and subcultured. Harvest two kinds of virus culture fluids respectively, concentrate, inactivate and purify to obtain two kinds of virus stock solutions after removing impurities; two kinds of virus stock solutions are sterilized, mixed, and adsorbed with adjuvants to prepare dual inactivated vaccines for hand, foot and mouth disease. The two-combined hand, foot and mouth disease inactivated vaccine of the invention has strong immunogenicity and no cross-inhibition effect between the two viruses, and can effectively prevent the current domestic hand, foot and mouth disease. The two-combined hand, foot and mouth disease inactivated vaccine of the invention has simple preparation process and low production cost, and is suitable for large-scale production of vaccines.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a vaccine and a preparation method thereof. Background technique [0002] Hand foot and mouth disease (HFMD) is an infectious disease in children, also known as exanthematous vesicular stomatitis. It mostly occurs in children under the age of 5, and can cause herpes on the hands, feet, and oral cavity. The main clinical symptoms of the disease are herpes or ulcers on the mucous membranes of the hands, feet, and oral cavity. A small number of children can cause myocarditis, pulmonary edema, aseptic meningoencephalitis and other complications. In some severely ill children, if the disease develops rapidly, it will lead to death. [0003] Hand, foot and mouth disease is an infectious disease caused by enteroviruses. Enteroviruses are highly contagious, spread widely, and spread quickly. The epidemic of the disease can occur throughout the year, and has the characteristics of periodic ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N7/00A61K39/295A61K39/125A61P31/14C12R1/93
Inventor 于晓方张文艳李景良
Owner JILIN UNIV
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