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A combined vaccine against hand, foot and mouth disease

A combined vaccine and virus technology, applied in the direction of vaccines, multivalent vaccines, viruses, etc., can solve the problems of affecting the immune effect, unknown immune interference, reducing the immunogenicity of vaccine single components, etc., to achieve simplified vaccination procedures and consistent processes Good performance and improved inoculation efficiency

Active Publication Date: 2021-01-22
SINOVAC BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, there is no relevant report on the development of a combined vaccine of EV71, CA16 and CB3. First, a large-scale outbreak of hand, foot and mouth disease caused by EV71, CA16 and CB3 virus has occurred in a relatively short period of time. Secondly, hand, foot and mouth disease caused by virus There is a certain regional distribution. For developed countries with a relatively low incidence, there are few institutions investing in research and development. Third, the immune interference between different viral antigens is unknown, and this interference may directly reduce the efficacy of single-component vaccines. Immunogenicity, affecting immune effect

Method used

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  • A combined vaccine against hand, foot and mouth disease
  • A combined vaccine against hand, foot and mouth disease
  • A combined vaccine against hand, foot and mouth disease

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0045] The preparation of embodiment 1 virus liquid

[0046] 1. Cell culture and recover SLF-1 cells, use MEM solution containing 20% ​​bovine serum, culture at 37±1°C, replace the medium after adherence, and use MEM solution containing 10% bovine serum. Wash the monolayer, digest with 0.25% trypsin, and divide into culture flasks or cell factories at a ratio of 1:4, transfer to 40-layer cell factories, grow to a monolayer, and prepare for inoculation of viruses.

[0047] 2. Virus culture

[0048] The cells were inoculated with EV71 virus at an MOI of 0.0001, cultured at 36±1°C with a culture medium containing 2% bovine serum, and the virus was harvested after 8 days.

[0049] The CA16 virus was inoculated into the cells at an MOI of 0.001, cultured at 35±1°C with a culture solution containing 1% bovine serum, and the virus was harvested after 10 days.

[0050] The CB3 virus was inoculated into the cells at MOI 0.001, cultured at 35±1°C with a culture medium containing 2% bo...

Embodiment 2

[0051] The inactivation of embodiment 2 virus

[0052] (1) Clarification: Filter the virus liquid through a filter element with a continuous two-stage pore size of 3-0.8 μm and 0.65-0.22 μm or centrifuge (including continuous flow centrifugation) with a rotation speed of 3000 rpm and a centrifugation time of 0.5 hours to obtain a clarified virus liquid;

[0053] (2) Inactivation Divide EV71, CA16, and CB3 virus clarified liquid into 9 groups respectively, and add different volumes of 1:200 formalin solution to each group so that the theoretical final concentration is 200, 100, 67 μg / ml, each Concentrations of 3 groups of parallel samples were inactivated at 37±1°C. See Table 1 for grouping information. Sampling was carried out at 0, 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, and 24 hours of inactivation, and immediately after the sampling was completed, formaldehyde was neutralized with sodium bisulfite solution, and virus titration was carried out. degree measurement. The viru...

Embodiment 3

[0063] The purification of embodiment 3 virus

[0064] (1) Ultrafiltration concentration of virus liquid Select ultrafiltration membrane packs with molecular weight cut-offs of 100kD, 300kD and 500kD three pore sizes, respectively carry out ultrafiltration concentration of EV71 (or CA16 or CB3) with the same batch of inactivation liquid, and fix the ultrafiltration pressure , concentrate the same multiple (100 times), fix the number of times of diafiltration (8 times of diafiltration), compare the ultrafiltration time and the antigen recovery rate of three membrane packs, the removal rate of impurity protein and the activity ratio of the product before and after ultrafiltration concentration (antigen content / protein content), and detect the antigen content of each filtrate. Determine the pore size of the ultrafiltration membrane package according to the test results. The results are shown in Table 4.

[0065] Table 4 Comparison results of ultrafiltration effects with differ...

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Abstract

The invention provides a combined vaccine for preventing hand-foot-and-mouth disease. The combined vaccine contains an inactivated EV71 antigen, CA16 antigen and CB3 antigen. Main pandemic virus strains such as EV71, CA16 and CB3 causing the hand-foot-and-mouth disease are cultivated and subjected to ultrafiltration, concentration, sucrose density gradient centrifugation, ultrafiltration desugarization and sterile filtration, a purified virus liquid is obtained, and the combined vaccine which is used for preventing the hand-foot-and-mouth disease and has good immunogenicity and high safety and stability is further prepared from the virus liquid. The vaccine can prevent bodies from being infected by various enteroviruses, and the antigens are not interfered, so that the corresponding immunogenicity cannot be reduced compared with the immunogenicity stimulated by a single antigen; moreover, with the adoption of the vaccine, the vaccination procedure can be remarkably simplified, the vaccination efficiency is improved, and the cost is reduced.

Description

technical field [0001] The invention relates to the technical field of preparation of biological products, in particular to a combined vaccine containing enterovirus antigens for preventing hand, foot and mouth disease. Background technique [0002] Hand-foot-mouth disease (HFMD) is listed as a Class C infectious disease in my country's "Infectious Disease Prevention and Control Law". It is an acute infectious disease caused by a variety of enteroviruses, and it is more common in infants and young children. . Patients manifested as mouth pain, anorexia, low fever, small herpes or small ulcers on the hands, feet, mouth and other parts. Most patients healed themselves in about a week, and a few patients could cause myocarditis, pulmonary edema, aseptic meningoencephalitis, etc. complication. Individual critically ill children develop rapidly and lead to death. In addition to the more common types of EV71 (Enterovirus 71, EV71) and CA16 (Coxasckievirus A16, CA16), viruses tha...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A61K39/125A61K39/39A61P31/14
CPCA61K39/12A61K39/39A61K2039/5252A61K2039/55505A61K2039/70C12N2770/32034C12N2770/32334
Inventor 张燕金亚明刘可心李雅静张星星高强尹卫东
Owner SINOVAC BIOTECH
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