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Bivalent vaccine for hemorrhagic fever with renal syndrome and its preparation method

A technology for renal syndrome and hemorrhagic fever, applied in biochemical equipment and methods, microbe-based methods, microbes, etc., can solve the problem of underrepresentation of vaccine strains

Inactive Publication Date: 2014-02-05
WUHAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

It can be shown that the representativeness of these vaccine strains is not strong. Therefore, to control the prevalence of HFRS, it is necessary to have sufficient understanding of the law and nature of the prevalence and evolution of HFRS. Vaccines that are more targeted and preventive

Method used

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  • Bivalent vaccine for hemorrhagic fever with renal syndrome and its preparation method
  • Bivalent vaccine for hemorrhagic fever with renal syndrome and its preparation method
  • Bivalent vaccine for hemorrhagic fever with renal syndrome and its preparation method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0020] Embodiment 1 Obtaining and identification of Hantavirus (Hantavirus) HV004

[0021] 1. Virus collection and sequencing

[0022] (1) Rat lung tissue, mite vector suspension, and blood samples from HFRS patients were collected from the middle and lower reaches of the Yangtze River from 1985 to 2009.

[0023] (2) Indirect immunofluorescence (IFA), enzyme-linked immunosorbent assay (EILSA), (Reverse transcriptase- Polymerase chain reaction, RT-PCR) method detection, screening mouse, mite, human virus antigen positive samples. The RT-PCR screening process is as follows: use the virus-specific RNA extraction kit (Promega) to extract the total RNA of the sample; detect the RNA content with a nucleic acid analyzer, and the RNA OD260 / 280 of all samples is ≥ 1.9, and then take 50ng RNA for RT-PCR reaction; Random primers were used for reverse transcription, and the reaction conditions were 37°C for 60 minutes, 93°C for 5 minutes to stop the reaction, and then quickly cooled on ...

Embodiment 2

[0033] Example 2 Preparation of virus vaccine

[0034] 1. Preparation of monovalent vaccine

[0035] (1) Continuously pass HV004 through the brain tissue of BALB / C suckling mice for 8 generations, harvest the brains of the affected mice, grind them into a homogenate, add protamine sulfate (final concentration: 1mg / ml), and initially remove tissue DNA and exogenous DNA The content is not more than 10ng / dose.

[0036] (2) After centrifugation at 110,000 g for 2.5 hours in a sucrose density gradient zone, a bright band between 30% and 45% and between 45% and 60% was obtained to purify the virus particles. Virus titer was 6.5lgCCID50 / ml; bacteria test and mycoplasma test were negative, and the virus titer was 1:1024 by IFA method.

[0037] (3) Use 2% V / V HEPES 1mol / L pH7.5 and 0.14% W / V NaHCO3 to adjust the pH of the purified virus stock solution to 7.4 at 4°C, and add 10% W / V BPL dropwise to make the final concentration 1 / 4000. After 24 hours of inactivation, the inactivated...

Embodiment 3

[0047] Embodiment 3 Animal experiments

[0048] The following is a detailed description of the monovalent vaccine as an example:

[0049] 1. Vaccine safety experiment

[0050] (1) Experimental research on animal allergy

[0051] 10ml of the sample was taken, and three guinea pigs were subcutaneously inoculated with calf serum as a control, and the guinea pigs were selected between 250-350g. Each animal was inoculated with 1ml, and the second injection was performed one week later, and each guinea pig was sensitized with 1ml subcutaneously. After three weeks of sensitization, each guinea pig was intravenously injected with the same sample 0.5ml, and no allergic reaction was observed in guinea pigs.

[0052] (2) Abnormal toxicity test

[0053] For the abnormal toxicity test, two kinds of animal tests, Balb / C and guinea pig, should be carried out, and the test animals should reach the clean level. A mouse with a body weight of 18-22 g is selected, and a guinea pig is selected...

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Abstract

The invention discloses a vaccine for hemorrhagic fever with renal syndrome (HFRS) and its preparation method. The vaccine is an inactivated virus of Hantavirus HV004 strain, and the virus has a preservation number of: CCTCC No. v201139. The inactivated virus can effectively prevent and treat HFRS. In addition, the invention also discloses a bivalent vaccine consisting of inactivated virus HV004 and Hantavirus Hubei-I strains. Experiments show that the bivalent vaccine has significant effects in prevention and treatment of HFRS. The invention also provides a preparation method of the vaccine, and the method includes virus proliferation, inactivation, purification and other steps. The bivalent vaccine of the invention provides an effective approach to prevention and treatment of HFRS, and plays an important role in controlling prevalence of the disease.

Description

technical field [0001] The invention relates to a novel Hantavirus and a bivalent vaccine for hemorrhagic fever with renal syndrome prepared from the virus, and also relates to a preparation method of the vaccine. Background technique [0002] Hemorrhagic fever with renal syndrome (HFRS) is a natural foci disease caused by Hantaviruses and transmitted by rodents. It has regional, explosive, periodic and variable epidemics. features. The HFRS epidemic area in my country has expanded from more than 600 counties and cities in the 1980s to more than 1300 counties and cities now, and the types of epidemic areas have also changed; in foreign countries, new-type epidemic areas are also expanding and evolving. People get sick by infecting viruses through aerosol inhalation, wounds, digestive tract and other ways. The disease has a high mortality rate and is very harmful. my country is the most severely affected country by hantavirus infection. In the past ten years, 25,000 to 60,...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N7/00A61K39/12A61P31/14C12R1/93
Inventor 杨占秋凌佳馨李金林鲁力张一卉刘东瀛刘媛媛罗凡熊海蓉侯炜肖红王继麟张云
Owner WUHAN UNIV
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