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Tetravalent dengue virus subunit vaccine, its preparation method and application thereof

A dengue virus, vaccine technology, applied in antiviral agents, viral antigen components, resistance to vector-borne diseases, etc.

Inactive Publication Date: 2012-12-19
MICROBE EPIDEMIC DISEASE INST OF PLA MILITARY MEDICAL ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although the attenuated live vaccines developed by Mahidol University in Thailand and the Walter Reed Army Research Institute in the United States have been clinically tested for many years by traditional cell passage methods, one or more serotypes in the quadrivalent vaccine mixture will enhance or inhibit Replication and immunogenicity of other serotypes in the vaccine, and ADE cannot be avoided

Method used

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  • Tetravalent dengue virus subunit vaccine, its preparation method and application thereof
  • Tetravalent dengue virus subunit vaccine, its preparation method and application thereof
  • Tetravalent dengue virus subunit vaccine, its preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0049] Embodiment 1, the preparation of four kinds of albumen

[0050] 1. E structural protein domain III protein of dengue virus DENV1

[0051] 1. Gene

[0052] Use dengue type 1 virus GZ / 80 strain RNA (the virus sequence has the 1st-10735th nucleotide sequence of GENBANK number AF350498) as a template, or use the artificially synthesized gene shown in SEQ ID NO: 5 as Template, PCR amplification was carried out with the primers shown in DIII1-F / DIII1-R in Table 1.

[0053] The PCR cycle conditions were 94°C for 2min; 94°C for 30sec, 55°C for 30sec, and 72°C for 1min, a total of 35 cycles; 72°C for 7min.

[0054] As a result, an amplified fragment with a length of about 300 bp was obtained, which was consistent with the gene length of the DIII region of E protein.

[0055] 2. Recombinant expression vector

[0056] The PCR amplified product obtained in step 1 was digested with EcoR V and Sal I at 37°C for 3 hours, and the purified fragment was recovered and ligated with the...

Embodiment 2

[0084] Embodiment 2, animal immunization and immune effect evaluation

[0085]Four-week-old Balb / C female mice were selected for immunization. Mice were divided into 2 groups (DIII mixed group and Trx group), 6 mice / group. Mix the purified DIII proteins of the 4 serotypes of dengue virus in a mass ratio of 1:1:1:1 (the immunization dose is 25 μg for the 4 proteins respectively) or dilute 25 μg of thioephrin (Trx) in 100 μl of PBS , and then mixed with Freund's complete adjuvant at a volume ratio of 1:1, fully emulsified and injected subcutaneously at multiple points for primary immunization, and each mouse was injected with 200 μl. Booster immunization once every 2 weeks, a total of 3 booster immunizations. The dosage and immunization route of each booster immunization were the same as those of the initial immunization, and the protein and Freund's incomplete adjuvant were mixed at a volume ratio of 1:1 during the booster immunization. Before the first immunization and two ...

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Abstract

The invention discloses a tetravalent dengue virus subunit vaccine, its preparation method and an application thereof. The active component of a product for preventing and / or treating dengue virus is a composition composed of a protein as shown in a domain III of an envelope protein E of dengue virus DENV1, a protein as shown in a domain III of an envelope protein E of dengue virus DENV2, a protein as shown in a domain III of an envelope protein E of dengue virus DENV3 and a protein as shown in a domain III of an envelope protein E of dengue virus DENV4. It proves through experiments that by the use of the vaccine provided by the invention for immunization of body, humoral immune responses of body can be excited, neonatal rat infected with the four serotype dengue viruses can be protected and infection of the four serotype dengue viruses can be resisted. The invention is of important realistic significance for preventing infection of human dengue viruses.

Description

technical field [0001] The invention relates to a tetravalent dengue virus subunit vaccine and its preparation method and application. Background technique [0002] Dengue virus is an important member of the Flaviviridae family, which is widely distributed in tropical and subtropical regions, and its main vector is Aedes aegypti. The disease caused by this virus infection is one of the most important arbovirus diseases. According to WHO statistics, about 100 million people in the world are infected with dengue virus every year. With the global warming and the increase of international communication, the epidemic area of ​​the disease is expanding and has become a serious public health problem. my country's southeast coastal areas such as Guangdong, Fujian, Hainan and Taiwan are high-incidence areas for dengue fever. Since 1978, there have been dozens of outbreaks of dengue fever in this area. In 1980 and 1985 alone, the number of dengue fever outbreaks in Hainan exceeded 5...

Claims

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Application Information

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IPC IPC(8): A61K39/12A61P31/14
CPCY02A50/30
Inventor 秦成峰赵慧邓永强姜涛秦鄂德
Owner MICROBE EPIDEMIC DISEASE INST OF PLA MILITARY MEDICAL ACAD OF SCI
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