CN106699880A - Preparation method of egg yolk antibody for treating type I and novel duck hepatitis and duck glossocele - Google Patents
Preparation method of egg yolk antibody for treating type I and novel duck hepatitis and duck glossocele Download PDFInfo
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- CN106699880A CN106699880A CN201611182581.0A CN201611182581A CN106699880A CN 106699880 A CN106699880 A CN 106699880A CN 201611182581 A CN201611182581 A CN 201611182581A CN 106699880 A CN106699880 A CN 106699880A
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- 238000002360 preparation method Methods 0.000 title claims abstract description 16
- 108010000912 Egg Proteins Proteins 0.000 title abstract description 11
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Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K16/00—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies
- C07K16/08—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from viruses
- C07K16/10—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from viruses from RNA viruses
- C07K16/1009—Picornaviridae, e.g. hepatitis A virus
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K16/00—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies
- C07K16/02—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies from eggs
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K16/00—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies
- C07K16/08—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from viruses
- C07K16/081—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from viruses from DNA viruses
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K39/00—Medicinal preparations containing antigens or antibodies
- A61K2039/505—Medicinal preparations containing antigens or antibodies comprising antibodies
- A61K2039/507—Comprising a combination of two or more separate antibodies
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
- Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change
Abstract
The invention provides a preparation method of an egg yolk antibody for treating type I and novel duck hepatitis and duck glossocele. According to the technical scheme, a type I inactivated vaccine for duck hepatitis, a novel inactivated vaccine for the duck hepatitis and an inactivated vaccine for the duck glossocele are taken as immunogens for inoculating laying hen, and an immunization method and an inoculation dosage are designed in an innovative way, so that each antibody reaches a high titer level. On the basis, the extraction opportunity of the egg yolk antibody is determined by taking the specific titer indexes of different antibodies, so that the treatment effect of the obtained egg yolk antibody is ensured effectively. Meanwhile, the invention further designs a specific egg yolk antibody extraction process. The obtained triple egg yolk antibody has a high titer, and has definite prevention and treatment effects on the duck hepatitis and the duck glossocele at the same time; the method fills the technical blank of treatment of the duck glossocele in the field of egg yolk antibodies, and has a great popularization prospect; meanwhile, simultaneous prevention and treatment of the two diseases is realized.
Description
Technical field
The present invention relates to technical field of biological product preparation, while being related to aquatic bird prevention from suffering from the diseases and Control Technology, specifically relate to
And a kind of Yolk antibody preparation method for treating I types, new duck hepatitis and duck glossocele.
Background technology
Duck virus hepatitis (Duck Viral Hepatitis, DVH) is a kind of acute height lethal for endangering duckling
Infectious disease, is characterized in morbidity is anxious, the course of disease is short, propagation is fast, case fatality rate is high etc., mainly the duckling in 3 week old of infringement, and adult duck can
Infect but do not fall ill.After duckling morbidity, death steeply rises, dead after 5d to stop；Morbidity age in days is smaller, and case fatality rate is got over
Height, generally more than 30%, person high is up to 95% or even 100%.Die of illness in opisthotonos sample outward appearance more than duck, its liver often has spy
Levying property hemorrhagic lesions.
The cause of disease of duck hepatitis includes I types, II type type III row DHVs.Wherein in China's Major Epidemic I types.It is near
Year, I types occurred in that variant, have scholar by I type DHVs according to the phylogenetic analysis country in China and surrounding countries
Different strains are divided into 3 genotype i.e. A types, Type B and c-type, and the former tradition serum I type of A types correspondence, Type B correspondence Taiwan is new, c-type
Correspondence Korean Utility.The currently a popular DHV strains of China are main based on A types and c-type, but without cross protection between 2 serotypes
Power, but the duck hepatitis vaccine in China market is primarily directed to A type strains, thus it is speculated that it is probably to cause China to duck viral
One of the reason for hepatitis can not carry out effective prevention and control.
" glossocele ", in the second half year in 2014 Jiangsu start occur, afterwards again spread to Shandong, Henan, Hebei, Anhui,
The whole nation such as Jiangxi multiple duck culturing area.Suffer from duck because beak development causes that tongue is protruded outside, so as to gain the name slowly.Sick duck earliest can be
There is the symptom of tongue protrusion in 6 ages in days, and morbidity later stage beak is shorter than normal duck by 10%~30%, and once morbidity is difficult to recover just
Often.Cut open inspection sick duck finds, in addition to intestinal inflammatory, without obvious lesion.The disease just takes place in commercial generation Cherry Village Duckss,
Involved meat duck and kind duck later.Studies in China personnel have proven to the disease to be caused by parvovirus, and is referred to as " the short beak of duck
Syndrome fond of gossip " or " the short beak of duck and runting syndrome ".Because tongue is protruded, sick duck is caused normally to search for food, therefore sick duck
Often performance depauperation, leg, wing are easily broken, and such residual duck is rejected in many slaughterhouses so that raiser's loss is serious.At present, should
Disease has the trend for growing in intensity, it is necessary to cause enough attention.
It has been reported that, the strain that duck " glossocele " illness duck is separate is identified with various common duck virosis specific primer PCRs
Afterwards, only goose parvovirus is the positive.Found by viral key VP3 gene sequencing and comparison, VP3 bases of " glossocele " virus
Because sequence and " gosling plague " virus are nearest goose parvovirus (GPV) homology, 93.1%~98.2% is reached；It is tiny with kind duck
Viral (MDPV) homology is 81.5%~91.4%.The VP3 phylogenetic analysis of drafting display that " the big tongue being separated to
Disease " virus recently, has document to be temporarily referred to as Duck parvovirus (duck with two plants of genetic affinities of gosling plague Taiwan separation strains
parvovirus,DPV)。
At present, Yolk antibody patent relevant with duck hepatitis is concentrated mainly on extraction (" one kind preventing and treating duckling of Yolk antibody
The duck Yolk antibody and preparation method of virus hepatitis ", number of patent application 201110130086.6), the formulation of Yolk antibody
(" duck virus hepatitis duck plague bigeminy yolk antibody oral liquid and preparation method thereof ", number of patent application 201410762718.4) etc.
Aspect, the Yolk antibody patent to duck " glossocele " not yet finds at present.
The content of the invention
It is contemplated that for the technological deficiency of prior art, there is provided one kind treats I types, new duck hepatitis and duck glossocele
Yolk antibody preparation method, with solve in the prior art for duck glossocele epidemic method effect on driving birds is not good technical problem.
Another technical problem to be solved by the present invention is that lacking a kind of effective duck virus hepatitis yolk in the prior art
Antibody.
The invention solves the problems that another technical problem be to be difficult to duck based on the immunization method of Yolk antibody in the prior art
Virus hepatitis and duck glossocele realize preventive effect simultaneously.
To realize above technical purpose, the present invention uses following technical scheme：
A kind of Yolk antibody preparation method for treating I types, new duck hepatitis and duck glossocele, comprises the following steps：
1) hen is taken, duck hepatitis I types inactivated vaccine, duck hepatitis inactivated vaccine, duck glossocele inactivation epidemic disease is utilized respectively
Seedling inoculation；
2) when duck hepatitis I type antibody neutralization titer >=1 in the hen serum:2048th, in duck hepatitis novel antibodies and imitate
Valency >=1:2048th, duck glossocele antibody fine jade expands potency >=1:When 64, high-immunity egg is collected；
3) take step 2) obtained by high-immunity egg extract Yolk antibody.
Preferably, step 1) in the method for inoculation be：Both duck hepatitis I types inactivated vaccine and new inactivated vaccine are same
When each fundamental immunity injection once, then each reinforced immunological injection twice, per minor tick three weeks, every kind of vaccine immunity 1ml/ every time
Only；Above-mentioned fundamental immunity injects one week after, and inoculation is performed using duck glossocele inactivated vaccine, and duck glossocele is utilized after three weeks
Inactivated vaccine performs secondary immunity injection, and three inoculations, per injection are performed using duck glossocele inactivated vaccine after three weeks
Vaccine dose for 1ml/ only.
Preferably, the duck glossocele inactivated vaccine is using the DPV-HB plants of preparation of duck glossocele virus.
Preferably, step 3) the extraction Yolk antibody, including following operation：
A) by 10~15min of soaking disinfection in the bromogeramine solution of high-immunity egg immersion 1%, dry standby；
B yolk) is collected；
C yolk) is broken up with colloid mill and is made yolk liquid, and 1:4 add acetate buffer, stir 60min, use sheet frame mistake
Filter coarse filtration；
D the octanoic acid that cleaner liquid adds 2~4%) is crossed, 60min, plate-frame filtering to clarification is stirred；
E) Yolk antibody is obtained final product with 0.22 μm of micro porous filtration is degerming.
Preferably, step B) the collection yolk is to take machinery to beat eggs, and separates and collect yolk, and remove yolk system
Band.
Preferably, step C) pH value of the acetate buffer is 5.2.
Preferably, when step 2) in any one antibody titer reduction by one titre when, take the vaccine corresponding to the antibody
The 4th time is carried out to be immunized.
Yolk antibody prepared by above can be used to prevent and treat duck hepatitis, while duck glossocele can also be prevented and be controlled
Treat, concrete application method is as follows.
(1) consumption is treated：≤ 21 ages in days, chest muscle injection 2ml/ is only；>=21 ages in days, chest muscle injection 3ml/ is only.Can during severe
Duplicate injection 2~3 times.
(2) consumption is promptly prevented：≤ 21 ages in days, chest muscle injection 1ml/ is only；>=21 ages in days, chest muscle injection 2ml/ is only.Big stream
It is multiplicable during row.
The invention provides a kind of Yolk antibody preparation method for treating I types, new duck hepatitis and duck glossocele, the technology
Scheme is inoculated with egg using duck hepatitis I types inactivated vaccine, duck hepatitis inactivated vaccine, duck glossocele inactivated vaccine as immunogene
Chicken, the method and dosage of inoculation that devise immunity inoculation of novelty so that each antibody has reached titer level higher.
On the basis of this, the present invention determines the extraction opportunity of Yolk antibody with the specific potency index of different antibodies as condition, effectively protects
The therapeutic effect of gained Yolk antibody is demonstrate,proved.At the same time, the present invention further devises targetedly Yolk antibody and extracts work
Skill, the Triple egg yolk antibody of gained not only has potency higher, at the same have to duck hepatitis, duck glossocele definite prevention and
The technological gap that Yolk antibody field is treated to duck glossocele has been filled up in therapeutic effect, the foundation of the method, while realizing
Prevented and treated while stating two kinds of diseases, great promotion prospect.
Specific embodiment
Specific embodiment of the invention will be below described in detail.In order to avoid excessive unnecessary details,
Be will not be described in detail to belonging to known structure or function in following examples.In addition to being defined, institute in following examples
Technology and scientific terminology have the identical meanings being commonly understood by with those skilled in the art of the invention.
Embodiment 1
1st, the preparation of Triple egg yolk antibody
(1) acquisition of high-immunity egg：Commercially available duck hepatitis I type inactivated vaccines are chosen, split antenatal hen carries out three times and is immunized, every time
Interval two weeks, every time immune 1ml/.When duck hepatitis I type antibody neutralization titer >=1:When 2048, start to collect high-immunity egg.
(2) preparation of duck hepatitis I type Yolk antibodies is refined：Disappear being soaked in the bromogeramine solution of high-immunity egg immersion 1%
Malicious 15min, dries standby；Take machinery to beat eggs, separate and collect yolk；After being emulsified with colloid mill, 1:4 add acetate salt buffer
Liquid (pH5.2), stirs 60min, is filtered with plate filter；The octanoic acid that cleaner liquid adds 2% is crossed, 60min, sheet frame is stirred vigorously
Filtering；Cross that 0.22 μm of micro porous filtration of cleaner liquid is degerming to be obtained final product.
(3) application process：Duck hepatitis epidemic situation in 2000 7 age in days cherry valley ducks, the anti-duck prepared using the above method there is
Hepatitis I type Yolk antibodies, chest muscle injection, 2ml/ is only.Epidemic situation is controlled, and cure rate is 70%, and separated identification finds, duck group
Part infection is duck hepatitis new virus during morbidity.There is duck " glossocele " again during this duck 14 week old of group, use duck " big tongue
The conventional medicines such as disease " Yolk antibody are treated, and the state of an illness is eased, and cure rate is 90%, and body weight is compared with other ducks group when delivering for sale
Average low 0.4kg or so.
2nd, the application and therapeutic evaluation of Triple egg yolk antibody
There is duck hepatitis epidemic situation and (have I types, new mixed infection situation, and " big with duck in 2000 7 age in days cherry valley ducks
Glossopathy "), using anti-duck hepatitis I types-duck hepatitis it is new-duck " glossocele " Triple egg yolk antibody, chest muscle injection, 2ml/ only.Epidemic situation
It is effectively controlled, cure rate reaches 93%.
The duck occurred in duck hepatitis chooses 2 groups of ducks group (totally 4000,7 ages in days) not fallen ill temporarily and is promptly prevented,
Chest muscle is injected, and 2ml/ is only.After duck epidemic situation is controlled, there is no the list of duck hepatitis and duck " glossocele " in this 4000 meat ducks
Solely infection or mixed infection, prevention ＆ protection rate has reached 100%.
Embodiment 2
A kind of Yolk antibody preparation method for treating I types, new duck hepatitis and duck glossocele, comprises the following steps：
1) hen is taken, duck hepatitis I types inactivated vaccine, duck hepatitis inactivated vaccine, duck glossocele inactivation epidemic disease is utilized respectively
Seedling inoculation；
2) when duck hepatitis I type antibody neutralization titer >=1 in the hen serum:2048th, in duck hepatitis novel antibodies and imitate
Valency >=1:2048th, duck glossocele antibody fine jade expands potency >=1:When 64, high-immunity egg is collected；
3) take step 2) obtained by high-immunity egg extract Yolk antibody.
On the basis of above technical scheme, following condition is met：
Step 1) in the method for inoculation be：Both duck hepatitis I types inactivated vaccine and new inactivated vaccine are while each basis
Once, then each reinforced immunological is injected twice for inoculation, every minor tick three weeks, and each every kind of vaccine immunity 1ml/ is only；It is above-mentioned
Fundamental immunity injects one week after, and inoculation is performed using duck glossocele inactivated vaccine, and epidemic disease is inactivated using duck glossocele after three weeks
Seedling performs secondary immunity injection, and three inoculations, the vaccine of per injection are performed using duck glossocele inactivated vaccine after three weeks
Dosage is 1ml/.
The duck glossocele inactivated vaccine is using the DPV-HB plants of preparation of duck glossocele virus.
Step 3) the extraction Yolk antibody, including following operation：
A) by 10~15min of soaking disinfection in the bromogeramine solution of high-immunity egg immersion 1%, dry standby；
B yolk) is collected；
C yolk) is broken up with colloid mill and is made yolk liquid, and 1:4 add acetate buffer, stir 60min, use sheet frame mistake
Filter coarse filtration；
D the octanoic acid that cleaner liquid adds 2~4%) is crossed, 60min, plate-frame filtering to clarification is stirred；
E) Yolk antibody is obtained final product with 0.22 μm of micro porous filtration is degerming.
Step B) the collection yolk is to take machinery to beat eggs, and separates and collect yolk, and remove chalaza.
Step C) acetate buffer pH value be 5.2.
When step 2) in any one antibody titer reduction by one titre when, taking the vaccine corresponding to the antibody is carried out the 4th time
It is immune.
Embodiment 3
A kind of Yolk antibody preparation method for treating I types, new duck hepatitis and duck glossocele, comprises the following steps：
1) hen is taken, duck hepatitis I types inactivated vaccine, duck hepatitis inactivated vaccine, duck glossocele inactivation epidemic disease is utilized respectively
Seedling inoculation；
2) when duck hepatitis I type antibody neutralization titer >=1 in the hen serum:2048th, in duck hepatitis novel antibodies and imitate
Valency >=1:2048th, duck glossocele antibody fine jade expands potency >=1:When 64, high-immunity egg is collected；
3) take step 2) obtained by high-immunity egg extract Yolk antibody.
Embodiments of the invention have been described in detail above, but the content is only presently preferred embodiments of the present invention,
It is not intended to limit the invention.All any modification, equivalent and improvement made in application range of the invention etc., all should
It is included within protection scope of the present invention.
Claims (7)
1. a kind of Yolk antibody preparation method for treating I types, new duck hepatitis and duck glossocele, it is characterised in that including following step
Suddenly：
1) hen is taken, duck hepatitis I types inactivated vaccine, duck hepatitis inactivated vaccine, duck glossocele inactivated vaccine is utilized respectively and is exempted from
Epidemic disease is injected；
2) when duck hepatitis I type antibody neutralization titer >=1 in the hen serum:2048th, duck hepatitis novel antibodies neutralization titer >=
1:2048th, duck glossocele antibody fine jade expands potency >=1:When 64, high-immunity egg is collected；
3) take step 2) obtained by high-immunity egg extract Yolk antibody.
2. preparation method according to claim 1, it is characterised in that step 1) in the method for inoculation be：Duck hepatitis I
Each fundamental immunity injection simultaneously of both type inactivated vaccine and new inactivated vaccine once, then inject twice, every time by each reinforced immunological
Interval three weeks, each every kind of vaccine immunity 1ml/ is only；Above-mentioned fundamental immunity injects one week after, is held using duck glossocele inactivated vaccine
Row inoculation, secondary immunity injection is performed after three weeks using duck glossocele inactivated vaccine, is inactivated using duck glossocele after three weeks
Vaccine performs three inoculations, and the vaccine dose of per injection is 1ml/.
3. preparation method according to claim 1, it is characterised in that the duck glossocele inactivated vaccine is using the big tongue of duck
The DPV-HB plants of preparation of sick virus.
4. preparation method according to claim 1, it is characterised in that step 3) the extraction Yolk antibody, including following behaviour
Make：
A) by 10~15min of soaking disinfection in the bromogeramine solution of high-immunity egg immersion 1%, dry standby；
B yolk) is collected；
C yolk) is broken up with colloid mill and is made yolk liquid, and 1:4 add acetate buffer, stir 60min, use plate filter
Coarse filtration；
D the octanoic acid that cleaner liquid adds 2~4%) is crossed, 60min, plate-frame filtering to clarification is stirred；
E) Yolk antibody is obtained final product with 0.22 μm of micro porous filtration is degerming.
5. preparation method according to claim 4, it is characterised in that step B) the collection yolk is to take machinery to beat eggs,
Yolk is separated and collected, and removes chalaza.
6. preparation method according to claim 4, it is characterised in that step C) pH value of the acetate buffer is
5.2。
7. preparation method according to claim 1, it is characterised in that：When step 2) in any one antibody titer reduction by one
During titre, take the vaccine corresponding to the antibody carry out the 4th time be immunized.
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| Application Number | Priority Date | Filing Date | Title |
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| CN201611182581.0A CN106699880A (en) | 2016-12-20 | 2016-12-20 | Preparation method of egg yolk antibody for treating type I and novel duck hepatitis and duck glossocele |
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| Application Number | Priority Date | Filing Date | Title |
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| CN201611182581.0A CN106699880A (en) | 2016-12-20 | 2016-12-20 | Preparation method of egg yolk antibody for treating type I and novel duck hepatitis and duck glossocele |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107739406A (en) * | 2017-09-30 | 2018-02-27 | 天津瑞普生物技术股份有限公司 | A kind of preparation method of Goose Parvovirus, Muscovy duck parvovirus bigeminy Yolk antibody |
| CN110938135A (en) * | 2019-11-22 | 2020-03-31 | 甘肃天祁生物科技有限公司 | Solid-liquid separation method for suspension in pregnant mare serum gonadotropin crude product extraction process |
Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN110938135A (en) * | 2019-11-22 | 2020-03-31 | 甘肃天祁生物科技有限公司 | Solid-liquid separation method for suspension in pregnant mare serum gonadotropin crude product extraction process |
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