CN110652583A - Bivalent inactivated antigen of swine foot-and-mouth disease and swine fever low virulent strain bigeminy vaccine - Google Patents
Bivalent inactivated antigen of swine foot-and-mouth disease and swine fever low virulent strain bigeminy vaccine Download PDFInfo
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- A61K39/00—Medicinal preparations containing antigens or antibodies
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- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
- A61P31/12—Antivirals
- A61P31/14—Antivirals for RNA viruses
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- A61K39/00—Medicinal preparations containing antigens or antibodies
- A61K2039/51—Medicinal preparations containing antigens or antibodies comprising whole cells, viruses or DNA/RNA
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- A61K39/00—Medicinal preparations containing antigens or antibodies
- A61K2039/55—Medicinal preparations containing antigens or antibodies characterised by the host/recipient, e.g. newborn with maternal antibodies
- A61K2039/552—Veterinary vaccine
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- A—HUMAN NECESSITIES
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- A61K39/00—Medicinal preparations containing antigens or antibodies
- A61K2039/70—Multivalent vaccine
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- C12N2770/00—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA ssRNA viruses positive-sense
- C12N2770/00011—Details
- C12N2770/24011—Flaviviridae
- C12N2770/24311—Pestivirus, e.g. bovine viral diarrhea virus
- C12N2770/24334—Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein
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- C12N2770/00011—Details
- C12N2770/32011—Picornaviridae
- C12N2770/32111—Aphthovirus, e.g. footandmouth disease virus
- C12N2770/32134—Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein
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Abstract
The invention discloses a bivalent inactivated antigen of swine foot-and-mouth disease and a swine fever low virulent strain bivalent vaccine, which is characterized in that: according to 20-50 parts of each bottle, 0.2-0.5 part of hog cholera antigen venom, 0.5-1.25 parts of foot-and-mouth disease O-type antigen, 0.5-1.25 parts of foot-and-mouth disease Asia I-type antigen and 1.8-4.5 parts of milk sucrose protective agent; the foot-and-mouth disease O type virus and Asian I type virus are respectively concentrated and purified by 300 times through 200 times, the hog cholera attenuated virus is concentrated by 10 times, most heterologous proteins, heat sources and inactivators are removed, side reactions are reduced during use, and the bivalent vaccine can improve the immune antibody of the hog cholera by more than 8.75 times compared with a single vaccine when being diluted along with use, so that a convenient condition for preventing two diseases through one injection is provided for a user.
Description
Technical Field
The invention relates to the field of veterinary biological products, in particular to a method for preparing bigeminy vaccine by porcine foot-and-mouth disease O-type and Asia I-type inactivated antigens and concentrated swine fever low virulent strains after high-power concentration and purification by a biological concentration technology.
Background
Hog cholera, commonly known as "rinderpest", is a hog cholera virus of the genus Pestivirus (Pestivirus) of the virus family draped. The pig disease has high infectious epidemic disease, is one of main infectious diseases threatening the pig industry, and is characterized by acute, septic change, bleeding of parenchymal organs, necrosis and infarction; the onset is mainly through direct contact, or due to contact with contaminating agents. The digestive tract, nasal mucosa and ruptured skin are all routes of infection.
The vaccines for preventing swine fever in the current market mainly comprise the following three vaccines:
1. classical swine fever live vaccine (I) -milk rabbit vaccine.
2. Classical swine fever live vaccine (II) -cell vaccine.
3. Classical swine fever live vaccine (I) -lymph spleen vaccine.
Preparation of the three vaccines:
1>preparing venom: preparing venom according to 2000 edition "veterinary biological product code" year swine fever live vaccine preparation and inspection code (I) method, wherein the virus titer is not less than 105RID/ML。
Vaccine preparation: calculated according to 20-50 parts of each bottle
Antigen 0.6-1.1ML, heat-resistant protective agent 0.9-1.6ML, subpackaging in 7ML glass bottles, and freeze-drying at 1.5-3ML per bottle.
Vaccine assay: and (3) executing the swine fever live vaccine manufacturing and inspection regulation (I), and storing the vaccine at the temperature of below 15 ℃ below zero for 18 months.
The using method comprises the following steps: the three vaccines were used in the same way, either by intramuscular or subcutaneous injection. When in use, the pig is diluted with the first part marked by a bottle label, and is diluted with 1ml of sterile physiological saline per first part, and the volume of the pig is 1ml for both big pigs and small pigs. The injection should be used up within 1 hour; if the temperature is 15-27 ℃, the catalyst should be used up within 2 hours. The injection is preferably at a time 2 hours after or before eating. The injection can generate immunity after 4 days, and the immune period can reach 12 months.
Foot-and-mouth disease is an acute fever and highly contagious infectious disease of artiodactyl caused by foot-and-mouth disease virus of picornaviridae. Clinically, it is characterized by blister and ulceration of oral mucosa, nasal osculum, hoof and breast skin of pig. The foot-and-mouth disease of pigs has high morbidity, fast infection and large epidemic scope, can cause mass death to piglets, causes serious economic loss, and has been an international key quarantine object because epidemic prevention is very important to the foot-and-mouth disease in all countries in the world.
At present, the inactivated vaccines for preventing the foot-and-mouth disease in the market mainly comprise the following two types:
inactivated vaccine for O-type foot-and-mouth disease of pig.
Bivalent inactivated vaccine for O-type and I-type foot-and-mouth diseases of pigs.
The preparation of the two vaccines:
1>preparing venom: preparing venom according to the preparation and inspection method of vaccine for pig foot and mouth disease with virus titer not less than 107LD50/0.2ML.
Vaccine preparation: inoculating O-type or Asia I-type foot-and-mouth disease virus to BHK21 cell culture, respectively harvesting infected cell fluid, removing cell debris from the virus fluid, inactivating by BEI, and adding blocking agent to terminate inactivation. Mixing the two antigens in equal amount, emulsifying the antigen 50% and oil adjuvant 50% to obtain milky white or light red viscous emulsion, packaging in 50 or 100ML plastic bottles, and storing at 2-8 deg.C.
Vaccine assay: executing the manufacturing and inspection rules of the inactivated vaccine for the foot-and-mouth disease of the pigs, and storing the qualified vaccine at 2-8 ℃ with the validity period of 1 year.
The using method comprises the following steps: the application method is the same, and the injection is administered by intramuscular injection after application of auricle, and the weight of each pig is 10-25kg, 1 mL; each pig with the weight of above 25kg is 2mL. It can be used for preventing O-type and I-type foot and mouth diseases of pig, and has immunity of 6 months after injection of vaccine 15 days.
Under the condition that both antigens have, the final vaccine preparation process is changed, and inactivated antigens of the foot-and-mouth disease type O and the Asia type I are respectively prepared, highly concentrated and purified by a biological concentration technology, and then freeze-dried with concentrated swine fever low virulent strains to form bigeminy vaccines. Thoroughly solves the phenomenon that the foot-and-mouth disease antigen is damaged in mineral oil for a long time, removes most heterologous proteins, heat sources and inactivators, ensures the effectiveness and stability of the antigen, concentrates the hog cholera attenuated virus by 10 times, removes the heterologous proteins and lightens side reactions; can improve the immune antibody of the swine fever by more than 8.75 times compared with the single vaccine, provides a convenient condition for preventing two diseases by one injection for users, and is a problem which needs to be solved urgently by research personnel in the field.
Disclosure of Invention
The invention aims to solve the problems and provides a preparation method of a bivalent inactivated antigen of swine foot-and-mouth disease and a swine fever low virulent strain, wherein the bivalent inactivated antigen and the swine fever low virulent strain are diluted when used, and the bivalent vaccine can improve the immune antibody of swine fever by more than 8.75 times compared with a single vaccine, thereby providing a convenient condition for preventing two diseases by one injection for a user.
The technical scheme of the invention is that the bivalent inactivated antigen of the pig foot-and-mouth disease and the swine fever low virulent strain are combined vaccine, which is characterized in that: according to 20-50 parts of each bottle, 0.2-0.5 part of hog cholera antigen venom, 0.5-1.25 parts of foot-and-mouth disease O-type antigen, 0.5-1.25 parts of foot-and-mouth disease Asia I-type antigen and 1.8-4.5 parts of milk sucrose protective agent.
The preparation method comprises the following steps of calculating 20-50 parts of each bottle:
step (1), sterilizing the milk and sucrose protective agent for 30 minutes at 116 ℃ under high pressure according to the proportion of the formula, cooling to 30 ℃, adding sterile swine fever antigen venom under the aseptic condition, and uniformly stirring;
and (2) adding the sterile foot-and-mouth disease O-type antigen into the solution obtained in the step (1) according to the formula proportion under the sterile condition, and stirring and uniformly mixing.
And (3) adding the sterile Asia I-type antigen of the foot-and-mouth disease into the solution obtained in the step (2) according to the formula proportion under the aseptic condition, and stirring and uniformly mixing.
And (4) subpackaging the veterinary vaccine obtained in the step (3) in 7-20ML glass bottles under the aseptic condition, wherein each glass bottle has 3-7.5ML, freeze-drying according to the method of the swine fever live vaccine manufacturing and inspection procedure (I), and storing the vaccine at the temperature below 15 ℃.
According to 20 parts of each bottle, the swine fever antigen venom is 0.2 part, the foot-and-mouth disease O-type antigen is 0.5 part, the foot-and-mouth disease Asia I-type antigen is 0.5 part, and the milk sucrose protective agent is 1.8 parts.
According to 50 parts of each bottle, the swine fever antigen venom is 0.5 part, the foot-and-mouth disease O-type antigen is 1.25 parts, the foot-and-mouth disease Asian I-type antigen is 1.25 parts, and the milk sucrose protective agent is 4.5 parts.
Preferably, the swine fever antigen venom is prepared by a method of swine fever live vaccine manufacturing and inspection regulation (I) of 2000 edition veterinary biologics regulation, and is concentrated by 10 times, and the virus titer is not less than 5X105RID/ML。
Preferably, the foot-and-mouth disease O type virus titer and Asia I type virus titer are not less than 107LD500.2ML. was purified by 200-fold and 300-fold concentration respectively, so that the final antigen content, i.e. 146S, was 500ug/ML.
Compared with the prior art, the invention has the beneficial effects that:
in the bivalent vaccine obtained by the invention, two foot-and-mouth disease inactivated antigens are highly concentrated and purified by a biological concentration technology, and most of heterologous proteins, heat sources and residual inactivators are removed. All are stored below 15 ℃ below zero, the phenomenon that the foot-and-mouth disease antigen is damaged in mineral oil for a long time is thoroughly solved, the effectiveness and stability of the antigen are ensured, and the immune effect is better. Carrying out 10 times concentration on the hog cholera attenuated virus, removing heterologous protein and reducing side reaction. After the bivalent vaccine is immunized, the manufacturing and inspection regulation inspection of the inactivated vaccine for the foot-and-mouth disease is carried out, and the O-type and Asia-I-type antibodies of the foot-and-mouth disease meet the standard of using bivalent vaccine, so that the immune antibody of the swine fever is improved by more than 8.75 times compared with the immune antibody using a single vaccine. The period of the foot-and-mouth disease validity is prolonged from 12 months to 18 months.
All raw materials in the formula are easy to obtain and are produced by professional veterinary biological product production enterprises, so that the preparation is easy and the preparation can be popularized and applied in a large area.
Compared with the traditional vaccine, the bigeminal vaccine in the prior art has the advantages of convenience in preparation, good stability, strong effect, easiness in storage, prevention of two diseases by one injection, convenience in use, wide application range and the like, and can be widely applied to the field of biological products for livestock.
Detailed Description
Example 1:
according to 20 parts per bottle, the swine fever antigen venom is 0.2ML, the foot-and-mouth disease O-type antigen is 0.5ML, the foot-and-mouth disease Asian I-type antigen is 0.5ML, and the milk sucrose protective agent is 1.8 ML.
The preparation method comprises the following steps of calculating 20 parts per bottle:
step (1), sterilizing the milk and sucrose protective agent for 30 minutes at 116 ℃ under high pressure according to the proportion of the formula, cooling to 30 ℃, adding sterile swine fever antigen venom under the aseptic condition, and uniformly stirring;
and (2) adding the sterile foot-and-mouth disease O-type antigen into the solution obtained in the step (1) according to the formula proportion under the sterile condition, and stirring and uniformly mixing.
And (3) adding the sterile Asia I-type antigen of the foot-and-mouth disease into the solution obtained in the step (2) according to the formula proportion under the aseptic condition, and stirring and uniformly mixing.
And (4) subpackaging the veterinary vaccine obtained in the step (3) in 7ML glass bottles with 3ML per bottle under the aseptic condition, and freeze-drying according to the method of the swine fever live vaccine manufacturing and inspection procedure (I), wherein the vaccine is stored at the temperature of below 15 ℃.
Example 2:
the vaccine comprises 50 parts per bottle of 0.5ML hog cholera antigen venom, 1.25ML foot-and-mouth disease O-type antigen, 1.25ML foot-and-mouth disease Asian I-type antigen, and 4.5ML milk sucrose protective agent.
The preparation method comprises the following steps of calculating 50 parts per bottle:
step (1), sterilizing the milk and sucrose protective agent for 30 minutes at 116 ℃ under high pressure according to the proportion of the formula, cooling to 30 ℃, adding sterile swine fever antigen venom under the aseptic condition, and uniformly stirring;
and (2) adding the sterile foot-and-mouth disease O-type antigen into the solution obtained in the step (1) according to the formula proportion under the sterile condition, and stirring and uniformly mixing.
And (3) adding the sterile Asia I-type antigen of the foot-and-mouth disease into the solution obtained in the step (2) according to the formula proportion under the aseptic condition, and stirring and uniformly mixing.
And (4) subpackaging the veterinary vaccine obtained in the step (3) in 20ML glass bottles with 7.5ML per bottle under the aseptic condition, and freeze-drying according to the method of the swine fever live vaccine manufacturing and inspection procedure (I), and storing the vaccine at the temperature of below 15 ℃ below zero.
Application method of bivalent inactivated antigen of foot-and-mouth disease and swine fever low virulent strain bivalent vaccine
Intramuscular injection, the first part is marked according to a bottle label and diluted with the first part when in use, the first part is diluted with 1ml of sterile physiological saline, then the diluted first part is absorbed into an oil adjuvant with the same amount by a syringe, the mixed solution is blown and beaten once by the syringe or directly shaken up to emulsify immediately, and more than 10 kg of the mixed solution is 1 ml. The injection should be used up within 1 hour; if the temperature is 15-27 ℃, the catalyst should be used up within 2 hours. The injection is preferably at a time 2 hours after or before eating. The swine fever can generate immunity after being injected for 4 days, the immune period can reach 12 months, the immunity can be generated 15 days after the foot-and-mouth disease O type injection, and the immune period is 6 months.
The invention relates to a bivalent inactivated antigen of swine foot-and-mouth disease and a swine fever attenuated vaccine for veterinary use, wherein one vaccine is live virus, the other vaccine is inactivated virus, the bivalent vaccine is originated at home and abroad, the inactivated antigens of foot-and-mouth disease O type and Asia I type are respectively subjected to high-power concentration and purification, most of heterologous proteins, heat sources and inactivating agents are removed, the rest inactivating agents of the vaccine are diluted after purification, the antigenicity of the swine fever attenuated virus is not influenced, the phenomenon that the foot-and-mouth disease antigen is damaged in mineral oil for a long time is thoroughly solved, the effectiveness and the stability of the antigen are ensured, the swine fever attenuated virus is subjected to 10-time concentration, the heterologous proteins are removed, side reactions are reduced, the bivalent vaccine is diluted when being used, and then the instant emulsification is an example at home and abroad, the bivalent vaccine can improve the immune antibody of the swine fever by more than 8.75 times compared with a single vaccine, and provides a convenient condition for preventing two diseases by one injection for.
The bigeminal vaccine of the invention has the advantages of convenient preparation, good stability, strong effect, easy preservation, convenient use, wide application and the like, and can be widely applied in the field of biological products for animals.
The technical solutions described above only represent the preferred technical solutions of the present invention, and some possible modifications to some parts of the technical solutions by those skilled in the art all represent the principles of the present invention, and fall within the protection scope of the present invention.
Claims (5)
1. Bivalent inactivated antigen of foot-and-mouth disease and swine fever low virulent strain bigeminy vaccine are characterized in that: according to 20-50 parts of each bottle, 0.2-0.5 part of hog cholera antigen venom, 0.5-1.25 parts of foot-and-mouth disease O-type antigen, 0.5-1.25 parts of foot-and-mouth disease Asia I-type antigen and 1.8-4.5 parts of milk sucrose protective agent.
The preparation method comprises the following steps of calculating 20-50 parts of each bottle:
step (1), sterilizing the milk and sucrose protective agent for 30 minutes at 116 ℃ under high pressure according to the proportion of the formula, cooling to 30 ℃, adding sterile swine fever antigen venom under the aseptic condition, and uniformly stirring;
step (2), adding the sterile foot-and-mouth disease O-type antigen into the solution obtained in the step (1) according to the formula proportion under the aseptic condition, and stirring and uniformly mixing;
step (3), adding the sterile Asia I-type antigen of the foot-and-mouth disease into the solution obtained in the step (2) according to the formula proportion under the aseptic condition, and stirring and uniformly mixing;
and (4) subpackaging the veterinary vaccine obtained in the step (3) in 7-20ML glass bottles under the aseptic condition, wherein each glass bottle has 3-7.5ML, freeze-drying according to the method of the swine fever live vaccine manufacturing and inspection procedure (I), and storing the vaccine at the temperature below 15 ℃.
2. The bivalent inactivated antigen of swine foot-and-mouth disease and the attenuated strain of swine fever according to claim 1, wherein the bivalent inactivated antigen of swine foot-and-mouth disease and the attenuated strain of swine fever are as follows: according to 20 parts of each bottle, the swine fever antigen venom is 0.2 part, the foot-and-mouth disease O-type antigen is 0.5 part, the foot-and-mouth disease Asia I-type antigen is 0.5 part, and the milk sucrose protective agent is 1.8 parts.
3. The bivalent inactivated antigen of swine foot-and-mouth disease and the attenuated strain of swine fever according to claim 1, wherein the bivalent inactivated antigen of swine foot-and-mouth disease and the attenuated strain of swine fever are as follows: according to 50 parts of each bottle, the swine fever antigen venom is 0.5 part, the foot-and-mouth disease O-type antigen is 1.25 parts, the foot-and-mouth disease Asian I-type antigen is 1.25 parts, and the milk sucrose protective agent is 4.5 parts.
4. The bivalent inactivated antigen of swine foot-and-mouth disease and the attenuated strain of swine fever of claims 1 to 3, which is characterized in that: the swine fever antigen venom is prepared by a method of manufacturing and testing regulation (I) of swine fever live vaccine carried in 2000 edition 'veterinary biological product regulation', and is concentrated by 10 times, and the virus titer is not lower than 5X105 RID/ML.
5. The bivalent inactivated antigen of swine foot-and-mouth disease and the attenuated strain of swine fever as claimed in claims 1 to 3, which is a bivalent vaccineCharacterized in that: the titer of foot-and-mouth disease type O virus and Asia type I virus is not less than 107LD50/0.2ML, respectively through 200-fold and 300-fold concentration and purification, the antigen content before the matched seedling, i.e. 146S, is 500ug/ML.
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| CN201810696907.4A CN110652583A (en) | 2018-06-29 | 2018-06-29 | Bivalent inactivated antigen of swine foot-and-mouth disease and swine fever low virulent strain bigeminy vaccine |
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