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Duck viral hepatitis bivalent yolk antibody, preparation method and application thereof

A technology of duck viral hepatitis and egg yolk antibody, which is applied in the direction of virus/bacteriophage, antiviral immunoglobulin, botanical equipment and methods, etc., can solve the problems of lack of effective control measures and lack of cross-protection for DVH, and achieve prevention and control Duck viral hepatitis, significant social benefits, and high safety effects

Inactive Publication Date: 2014-06-18
PU LIKE BIO ENG
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] The research results have shown that DHAV-1 and DHAV-3 are prevalent in many areas of my country at the same time, and there is a lack of cross protection between the two, and there is still a lack of effective control measures for DVH caused by DHAV-3

Method used

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  • Duck viral hepatitis bivalent yolk antibody, preparation method and application thereof
  • Duck viral hepatitis bivalent yolk antibody, preparation method and application thereof
  • Duck viral hepatitis bivalent yolk antibody, preparation method and application thereof

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preparation example Construction

[0041] The preparation method of the duck viral hepatitis bivalent egg yolk antibody adopts the following steps: breeding of duck hepatitis virus DHAV-1 and DHAV-3 seed virus, preparation of vaccine for immunization, immunization of laying hens, and extraction of egg yolk antibody. The selected DHAV-1 strain in the embodiment of the present invention is DRL-62 strain, and DHAV-3 strain is SD1 strain, and its specific method is:

[0042] 1. The source of the virus species is duck hepatitis virus DRL-62 strain (preserved by ATCC, deposit number VR-1313); SD1 strain was isolated by Pulaike Bioengineering Co., Ltd. (CCTCC deposit, deposit number: CCTCC NO.V201225).

[0043] 2. Propagation of virus seeds Dilute duck hepatitis virus DRL-62 strain virus seeds with sterile PBS solution at 1:100, inoculate 9-11-day-old SPF chicken embryos through the allantoic cavity, 0.1ml per embryo. Continue incubation at 37°C, discard dead embryos within 48 hours, harvest dead chicken embryos withi...

Embodiment 1D

[0061] Molecular biological identification of embodiment 1DHV SD1 isolate

[0062] 1. Duck hepatitis virus SD1 strain virus is a newly isolated and identified virus, which has the following characteristics:

[0063] 1.1 The clinical symptoms caused by DHAV-3 and DHAV-1 are very similar. The main manifestations are: the ducklings have sudden onset, obvious neurological symptoms, convulsions, and death soon. After death, the ducklings are in opisthotonus posture. The pathological changes showed that the liver was enlarged, and there were a large number of bleeding points and spots on the liver surface, and the bleeding became more obvious with the prolongation of the death time.

[0064] 1.2 According to the DHV genome sequence published in GenBank, several pairs of specific primers were designed, the VP1 gene was amplified by conventional RT-PCR method, and the sequence analysis of the VP1 gene of the isolate SD1 was carried out. The results showed that the VP1 gene of SD1 had...

Embodiment 2

[0096] Embodiment 2 Serum cross-neutralization test

[0097] The serum was fixed and diluted, and the positive serum of the DHV DRL-62 strain and the SD1 isolate were serially diluted 2 times with normal saline. Take 1.0ml of the virus solution of DRL-62 strain and SD1 isolate containing 200ELD50 / 0.2ml and mix them with equal amounts of positive sera of different dilutions of each strain, and act at 37°C for 1h. At the same time, virus and saline controls were set. Each neutralization group and control group were inoculated with 5 duck embryos through the allantoic cavity, 0.2m1 / embryo, cultured at 37°C, observed for 7 days, and recorded the number of dead embryos in each group. The neutralization titer of the serum was defined as the highest serum dilution for 50% duck embryo protection.

[0098] The results of the neutralization test showed that the isolated strain could be completely neutralized by the positive serum of the isolated strain, but could not be neutralized an...

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Abstract

The present invention provides a duck viral hepatitis bivalent yolk antibody and a preparation method thereof, wherein the bivalent yolk antibody comprises a duck viral hepatitis DHAV-1 type antibody and a duck viral hepatitis DHAV-3 type antibody. The preparation method comprises: (1) adopting a duck viral hepatitis DHAV-1 type strain and a duck viral hepatitis DHAV-3 type strain to respectively vaccinate SPF chicken embryo and susceptible duck embryo, harvesting allantoic fluid, mixing the harvested virus liquids according to a certain ratio, carrying out formaldehyde inactivation, and preparing a vaccine; (2) adopting the vaccine to immunize laying hens, sampling after immunization to determine whether the neutralizing titer of the anti-DHAV-1 type antigen antibody and the anti-DHAV-3 type antigen antibody in the chicken hyperimmune egg yolk is more than or equal to 1:8192, and collecting the hyperimmune egg of the chicken; and (3) disinfecting the eggshell of the hyperimmune egg, collecting the egg yolk, adding the equal volume of distilled water, uniformly stirring and mixing, carrying out low temperature pasteurization inactivation, adopting an acidification distilled water method to purify, adopting an octanoic acid method to purify, and carrying out micro-filtration and ultra-filtration. The duck viral hepatitis bivalent yolk antibody has characteristics of low cost and high titer, and can be provided for effectively controlling duck viral hepatitis caused by DHAV-1 and DHAV-3 so as to obtain significant social benefits.

Description

technical field [0001] The invention relates to a hyperimmune egg yolk antibody, in particular to an egg yolk antibody for duck viral hepatitis. Background technique [0002] Duck Virus Hepatitis (DVH) is an acute, highly fatal and severe infectious disease of ducklings caused by Duck Hepatitis Virus (DHV) of the family Picornaviridae Enterovirus. The disease occurs all the year round and only infects ducklings within 3 weeks of age in a natural state, often causing significant economic losses and threatening the healthy development of the duck industry. The World Organization for Animal Health (OIE) classifies duck viral hepatitis as a category B animal disease. [0003] Duck hepatitis virus has historically been divided into serotypes I, II and III, and there is no antigenic correlation among the three. Among them, type I duck viral hepatitis is distributed worldwide. Type II duck viral hepatitis was discovered in the United Kingdom in 1965, and since the outbreak in th...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N7/00C12N15/41C07K14/085C07K16/10C07K16/02A61K39/42A61P31/14A61P1/16C12R1/93
CPCA61K2039/505A61K2039/5252A61K2039/552C07K14/005C07K16/02C07K16/1009C07K2317/11C12N7/00C12N2770/32421C12N2770/32422C12N2770/32434
Inventor 张许科孙进忠白朝勇
Owner PU LIKE BIO ENG
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