阅读说明：本技术 一种快速筛选浆膜炎对型疫苗的方法 (Method for rapidly screening serositis paired vaccines ) 是由 孟照洁 翁立雪 许明清 刘丹 于 2019-10-25 设计创作，主要内容包括：本发明涉及一种快速筛选浆膜炎对型疫苗的方法,属于生物制品领域。包括疫苗筛选：选择浆膜炎疫苗；实验动物选择：选择雌兔,PCR检测大肠杆菌、沙门氏菌、鸭疫里氏杆菌、绿脓杆菌、李氏杆菌和巴氏杆菌,确定检测均为阴性；免疫：免疫选好的兔,采用背部、颈部多点免疫法,共选取6个点,进行三次免疫后大量收集血清冻干储备；疫苗筛选：将分离的鸭疫里氏杆菌加入生理盐水,制成浑浊液,煮沸1-2小时,与每个制备好的血清做玻片凝集。该方法收集全国合法浆膜炎灭活疫苗,兔作为免疫对象,使用不同厂家的浆膜炎灭活疫苗对其免疫,产生高免血清,保存血清,将临床鸭疫里氏菌与血清做玻片凝集,达到快速配型,筛选对型疫苗的方法。(The invention relates to a method for rapidly screening serositis paired vaccines, belonging to the field of biological products. The method comprises the following steps of vaccine screening: selecting a serositis vaccine; selecting experimental animals: selecting female rabbits, detecting escherichia coli, salmonella, riemerella anatipestifer, pseudomonas aeruginosa, listeria and pasteurella by PCR, and determining that the detection is negative; immunization: immunizing selected rabbits by a back and neck multi-point immunization method, selecting 6 points in total, performing three times of immunization, and collecting a large amount of serum for freeze-drying and storage; and (3) vaccine screening: adding separated Riemerella anatipestifer into normal saline to prepare turbid liquid, boiling for 1-2 hours, and performing glass slide agglutination with each prepared serum. The method collects national legal serositis inactivated vaccines, rabbits are used as immune objects, serositis inactivated vaccines of different manufacturers are used for immunizing the rabbits to generate hyperimmune serum, the serum is stored, clinical riemerella anatipestifer and the serum are subjected to slide agglutination to achieve rapid matching, and the matched vaccines are screened.)

1. A method for rapidly screening serositis paired vaccines is characterized in that: the method comprises the following steps:

1) and (3) vaccine screening: selecting all legal batch serositis vaccines;

2) selecting experimental animals: selecting female rabbits, and performing PCR detection on escherichia coli, salmonella, riemerella anatipestifer, pseudomonas aeruginosa, listeria and pasteurella to determine that the detection is negative, wherein the method can be used;

3) immunization: immunizing selected rabbits by a back and neck multi-point immunization method, selecting 6 points in total, performing three times of immunization, and collecting a large amount of serum for freeze-drying and storage;

4) and (3) vaccine screening: adding separated Riemerella anatipestifer into normal saline to prepare turbid liquid, boiling for 1-2 hours, performing glass slide agglutination with each prepared serum, and if agglutination occurs, selecting corresponding vaccine for immunization; if no agglutination occurs, the current vaccine can not form protection for the strain.

2. The method for rapidly screening serositis-paired vaccines according to claim 1, wherein in step 2), the experimental animals are selected from: female rabbits of 2-3kg were selected.

3. The method for rapidly screening serositis-paired vaccines according to claim 1, wherein in the step 3) immunization, the first immunization: each spot was immunized with 0.2ml, 1.2ml total, and a second immunization was performed after 21 days.

4. The method for rapidly screening serositis-paired vaccines according to claim 3, wherein in the step 3) immunization, the second immunization: each spot was immunized with 0.3ml, 1.8ml total, and a third immunization was performed after 21 days.

5. The method for rapidly screening serositis-paired vaccines according to claim 4, wherein in the step 3), the third immunization: each point was immunized with 0.5ml for a total of 3ml, and after 21 days of the third immunization, the serum was collected in large amounts and stored by lyophilization.

Technical Field

The invention belongs to the technical field of biological products, and particularly relates to a method for rapidly screening serositis vaccine.

Background

Riemerella anatipestifer is also called as infectious serositis of duck, and is a contact, acute or chronic infectious disease with septicemia. Is mainly characterized by cellulose pericarditis, cellulose perihepatitis, cellulose air sacculitis, caseous salpingitis, arthritis and paralysis.

The infectious serositis of duck is a chronic or acute septic infectious disease affecting ducks/geese caused by riemerella anatipestifer. Mainly infected ducks, geese, chickens and some wild birds can be infected. It is manifested as depression, anorexia, lacrimation, soft feet, thin and yellowish white or green feces, head-twisting and neck-circling, and intermittent convulsions until death. The pericardial fluid is increased or adhered to the epicardium, the liver is enlarged, or there are more cellulosic secretions on the surface. The morbidity mortality rate is generally 5% -80%, which often causes massive death of the ducklings/geese and leads to delayed development of the ducks/geese. It can be seen in four seasons, especially in autumn and winter. The traditional method for treating the infectious serositis of the duck is mainly based on medicines, seriously threatens the food safety and is released due to national banning of resistance. This has to be returned to vaccine prevention for the control of infectious serositis in ducks.

At present, the screening of the serositis vaccine is mainly evaluated by a method of attacking the vaccine after immunization, the operation is complex, the operation of test animals and professionals is needed, the screening period is at least 1 month, and the emergency immunity which is difficult to quickly prevent and control the disease is achieved.

Disclosure of Invention

In order to solve the technical problems, the invention provides a method for rapidly screening serositis paravaccine, which comprises the steps of immunizing rabbits with the purchasable national legal serositis vaccine to prepare hyperimmune serum, storing the serum, and once serositis occurs, performing slide agglutination on the separated Riemerella anatipestifer strain and the stored vaccine serum to achieve the effect of screening the paravaccine within 1 minute. The advantage of this patent is mainly convenient, swift, simple, has 1 month to shorten to 48 hours with the screening cycle, does not need professional test personnel and animal, and is economic high-efficient.

The invention is realized by the following technical scheme, and on one hand, the invention provides a method for rapidly screening serositis paired vaccines, which comprises the following steps:

1) and (3) vaccine screening: selecting all legal batch serositis vaccines;

2) selecting experimental animals: selecting female rabbits, and performing PCR detection on escherichia coli, salmonella, riemerella anatipestifer, pseudomonas aeruginosa, listeria and pasteurella to determine that the detection is negative, wherein the method can be used;

3) immunization: immunizing selected rabbits by a back and neck multi-point immunization method, selecting 6 points in total, performing three times of immunization, and collecting a large amount of serum for freeze-drying and storage;

4) and (3) vaccine screening: adding separated Riemerella anatipestifer into normal saline to prepare turbid liquid, boiling for 1-2 hours, performing glass slide agglutination with each prepared serum, and if agglutination occurs, selecting corresponding vaccine for immunization; if no agglutination occurs, the current vaccine can not form protection for the strain.

Preferably, in step 2), the experimental animals are selected: female rabbits of 2-3kg were selected.

Preferably, in the step 3) immunization, the first immunization: each spot was immunized with 0.2ml, 1.2ml total, and a second immunization was performed after 21 days.

Preferably, in the step 3) immunization, the second immunization: each spot was immunized with 0.3ml, 1.8ml total, and a third immunization was performed after 21 days.

Preferably, in the step 3) immunization, the third immunization: each point was immunized with 0.5ml for a total of 3ml, and after 21 days of the third immunization, the serum was collected in large amounts and stored by lyophilization.

The invention has the beneficial effects that:

1) the invention prepares hyperimmune serum by immunizing rabbits with a purchasable national legal serositis vaccine, stores the serum, and can agglutinate the separated riemerella anatipestifer strain and the stored vaccine serum by glass slides once the serositis occurs, thereby achieving the effect of screening out the vaccine against the type within 1 minute.

2) The advantage of this patent is mainly convenient, swift, simple, has 1 month to shorten to 48 hours with the screening cycle, does not need professional test personnel and animal, and is economic high-efficient.

Detailed Description

In order to make the objects, technical solutions and advantages of the present invention more apparent, the present invention is further described in detail with reference to the following embodiments. It should be understood that the specific embodiments described herein are merely illustrative of the invention and are not intended to limit the invention.

As introduced in the background art, currently, the screening of serositis-type vaccines is mainly evaluated by a method of attacking the vaccine after immunization, the operation is complex, experimental animals and professionals are required to operate, the screening period is at least 1 month, and the emergency immunization which can hardly realize quick prevention and control on the pathogenetic disease is realized. In order to solve the problems, the invention provides a method for rapidly screening serositis-associated vaccines.