CN102973933B - Bivalent vaccine for porcine reproductive and respiratory syndrome and classical swine fever prevention or treatment, and preparation method thereof - Google Patents
Bivalent vaccine for porcine reproductive and respiratory syndrome and classical swine fever prevention or treatment, and preparation method thereof Download PDFInfo
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Abstract
The present invention provides a bivalent vaccine for porcine reproductive and respiratory syndrome (PRRS) and classical swine fever (CSF) prevention or treatment. The bivalent vaccine comprises the following components: PRRS antigen with a content of at least 10<5>TCID50/unit, and CSF antigen with a content of at least 7500 rabbit infection dose/unit. The bivalent vaccine preparation method comprises: adopting passage cells to culture PRRS viruses and CSF viruses to obtain PRRS antigens and CSF antigens, mixing the two antigen components according to a certain ratio, and auxiliarily adopting a freeze drying protection agent to prepare the freeze drying vaccine. According to the present invention, the two antigens achieve the appropriate ratio, such that the bivalent vaccine achieves or exceeds efficacy of the original single vaccine; the bivalent vaccine has characteristics of simple preparation method, high titer content and convenient and rapid immunization; compared with fractionated immunization, vaccines for PRRS and CSF prevention and treatment through immunization at least 2 times or 3 times, and immunization methods thereof in the prior art, the bivalent vaccine and the immunization method thereof have characteristics of reduced immunization cost, saved immunization program, economy and reliability.
Description
Technical field
The present invention relates to bigeminy vaccine of a kind of prevention or treatment pig blue-ear disease and swine fever and preparation method thereof, belong to live vaccine field.
Background technology
Pig blue-ear disease claims again Porcine reproductive and respiratory syndrome (Porcine reproductive and respiratory syndrome, be called for short PRRS), be a kind of viral infectious that endangers pig industry, this disease is taking the respiratory tract disease of the breeding difficultys such as in-pig miscarriage, stillborn fetus, weak son, mummy tire and each age group pig and high mortality as feature.Summer and autumn in 2006, "swine high fever"of pig epidemic situation is sent out in south China part province, and the positive tissue scientific research personnel of the Ministry of Agriculture combines tackling key problem, determines that pig blue-ear disease variant viral is main pathogen, names as " highly pathogenic PRRS ".Until today, epidemic disease situation is still severe, the same with other viral diseases, there is no at present the treatment of effective curative drug for reproductive and respiratory syndrome, and immunity inoculation is this sick most effectual way of anti-system.
Swine fever (Classical Swine fever, be called for short CSF) be called again hog cholera (Hog cholera) or Europe class swine fever (European swine fever), it is the viral infectious disease of the acute or super febris acuta of a kind of pig, feature is clinically to disseminate rapidly, have a fever, and while dissecting inspection, can see typical hemorrhage pathological changes.Swine fever can infect the pig at various ages, popular throughout the year, and M & M is all high, very harmful to pig.Primary disease is to threaten one of most important infectious disease of pig industry.OIE (OIE) classifies swine fever one of as 16 kinds of Notifiable diseases of category-A, and China is decided to be a class deadly infectious disease.
At present, only have the univalent vaccine of pig blue-ear disease and swine fever, need separately injection respectively, not only immunization amount is large, and cost is very high, and repeatedly immunity causes larger stress to pig body, needs Immunity at intervals.But the bigeminy vaccine that there is not yet pig blue-ear disease and swine fever is successfully prepared all the time, it is anti-that bigeminy vaccine of the present invention can be realized a pin two, reduces immunostimulation, impels body can produce better immunne response.
Summary of the invention
In view of this, the bigeminy vaccine that main purpose of the present invention is to provide a kind of prevention or treatment pig blue-ear disease and swine fever, prevents simultaneously and treats pig blue-ear disease and hog cholera to realize.
Technical scheme
A bigeminy vaccine for prevention or treatment pig blue-ear disease and swine fever, comprises following composition: pig blue-ear disease antigenic component, swine fever antigen composition and vaccine freeze-drying protective agent.
Preferably, pig blue-ear disease antigenic content of the present invention is at least 10
5tCID
50/ head part, more preferably, pig blue-ear disease antigenic content of the present invention is 10
6tCID
50/ head part.
Preferably, swine fever antigen content of the present invention is at least 7500 rabbit infective doses/head part, and preferably, every part should be contained 0.015ml cell venom.
Preferably, swine fever antigen content of the present invention is 9000 rabbit infective doses/head part, and preferably, every part should be contained 0.01ml cell venom.
The preparation method of above-mentioned prevention or treatment pig blue-ear disease and swine fever bigeminy vaccine; comprise the following steps: use passage cell to cultivate PRRS virus and swine fever virus; to obtain pig blue-ear disease antigen and swine fever antigen; be mixed in proportion two kinds of antigenic components, be aided with freeze drying protectant and be prepared into freeze dried vaccine.
Preferably, in said method of the present invention, described pig blue-ear disease antigenic content is at least 10
5tCID
50/ head part;
Preferably, in said method of the present invention, described pig blue-ear disease antigenic content is at least 10
6tCID
50/ head part;
Preferably, in said method of the present invention, described swine fever antigen content is at least 7500 rabbit infective doses/head part.
Preferably, in said method of the present invention, described swine fever antigen content is 9000 rabbit infective doses/head part.
Preferably, in said method of the present invention, described antigen mixed volume ratio is pig blue-ear disease antigen: swine fever antigen volume ratio is 1: 5～1: 10.,, by being mixed in proportion two kinds of antigenic components, make the pig blue-ear disease antigenic content in bigeminy vaccine be at least 10
5tCID
50/ head part, more preferably, pig blue-ear disease antigenic content is 10
6tCID
50/ head part; Make swine fever antigen content be at least 7500 rabbit infective doses/head part, every part should be contained 0.015ml cell venom; Most preferably, swine fever antigen content is 9000 rabbit infective doses/head part, and every part should be contained 0.01ml cell venom.
Another object of the present invention is to provide the purposes of a kind of bigeminy vaccine as above aspect prevention or treatment pig blue-ear disease and swine fever.
Therefore, in bigeminy vaccine of the present invention, by making two kinds of antigens reach appropriate proportioning, two kinds of antigens are not disturbed mutually, vaccine potency at least reaches or exceeds the effect of original single Seedling.Secondly, bigeminy vaccine preparation method of the present invention is simple, the content of tiring of vaccine is high, immunity is convenient and swift, with gradation immunity of the prior art, at least need to make a call to 2 pins or 3 pins and could prevent and treat vaccine and the immunization method thereof of above two kinds of diseases and compare, reduce immune cost, save immune programme for children, reliable more economically.
Detailed description of the invention
Described in bigeminy vaccine of the present invention, PRRS virus is JXA1-R strain, is disclosed in Chinese patent CN 101307305A, and preserving number CGMCCNo.2467 is preserved in Chinese common micro-organisms culture presevation administrative center.
Swine fever virus described in bigeminy vaccine of the present invention is fever virus lapinized Chinese Strain, purchased from China Veterinery Drug Inspection Office, and bacterial strain deposit number CVCC AV1412.
In embodiment of the present invention bigeminy vaccine, preferably the front reproductive and respiratory syndrome virus content of lyophilizing is 10
9tCID
50/ ml; Before swine fever virus lyophilizing, content is 900,000 rabbit infective dose/ml.
In the embodiment of the present invention, can pass through to adjust the front reproductive and respiratory syndrome virus liquid of lyophilizing and swine fever virus liquid proportional to 1: 5～1: 10 volumes, can obtain the pig blue-ear disease antigenic content making in bigeminy vaccine and be at least 10
5tCID
50/ head part, more preferably, pig blue-ear disease antigenic content is 10
6tCID
50/ head part; Make swine fever antigen content be at least 7500 rabbit infective doses/head part, every part should be contained 0.015ml cell venom; Most preferably, swine fever antigen content is 9000 rabbit infective doses/head part, and every part should be contained 0.01ml cell venom.Wherein, in the embodiment of the present invention, preferably adjusting the front reproductive and respiratory syndrome virus of lyophilizing and swine fever virus ratio is 1: 5 volume ratio, and every part of bigeminy vaccine of acquisition is 10 containing reproductive and respiratory syndrome virus
6tCID
50, swine fever virus is 9000 rabbit infective doses.
The preparation of embodiment 1, pig blue-ear disease and swine fever bigeminal live vaccine
1. produce the preparation with seed culture of viruses
The preparation of pig blue-ear disease seed culture of viruses: get well-grown Marc-145 cell (purchased from China Veterinery Drug Inspection Office), with trypsinization, and be inoculated in cell bottle, seed culture of viruses is done to 10 times of dilutions by MEM (production of the Invitrogen company) maintenance medium containing 1%v/v hyclone (production of PAA company) simultaneously, by 5% volume inoculum concentration virus inoculation, 37 DEG C of cultivations, observation of cell pathological changes every day (CPE), in the time that appearring in 70% above cell, receives CPE poison, freeze thawing 3 times, results virus, virus titer is that every 1ml viral level is 10
9.5tCID
50.
The preparation of hog cholera seed culture of viruses: with α-MEM (production of the Invitrogen company) cell maintenance medium containing 1%v/v hyclone (production of PAA company), seed culture of viruses is made to the viral suspension of 0.3%v/v, inoculate well-grown ST cell (purchased from Chinese Typical Representative culture collection center) monolayer, put 37 DEG C and continue to cultivate.Change liquid once every results on the 2nd, get two receipts, three and receive cell culture venom as production seed culture of viruses.The every 1ml of cell seed culture of viruses is containing 900,000 rabbit infective doses of virus.
2. the cultivation of virus liquid preparation
The preparation of reproductive and respiratory syndrome virus liquid: get well-grown monolayer coverage rate and reach more than 80% Marc-145 cell, discard the MEM growth-promoting media containing 5%v/v hyclone, seed culture of viruses is done to 10 times of dilutions by the MEM maintenance medium containing 1%v/v hyclone simultaneously, press 5%v/v inoculum concentration virus inoculation, 37 DEG C of cultivations.Examine under a microscope the cytopathy (CPE) being caused by virus, until CPE harvesting culture while reaching 80%-90%, multigelation 3 times, results virus, virus titer is that every 1ml viral level is 10
9tCID
50.
The preparation of hog cholera venom: get the ST cell that forms good monolayer, inoculation, containing the maintenance medium (maintenance medium is the α-MEM culture fluid containing 1%v/v hyclone) of 3%v/v Cells for production seed culture of viruses, is put 37 DEG C and continued to cultivate.After connecing poison, within 3rd, do to gather in the crops and change liquid for the first time, change liquid once later every results on the 2nd, gather in the crops 3 times, the virus liquid of results is placed in-20 DEG C of following preservations.The every 1ml of virus liquid is containing 900,000 rabbit infective doses of virus.
3. viral level is measured
Reproductive and respiratory syndrome virus liquid viral level is measured: the 10 times of serial dilutions of MEM culture medium by the virus liquid of results with serum-free, get 10
-6, 10
-7, 10
-8, 10
-9, 10
-10, 10
-116 dilution factors, are inoculated in respectively 96 porocyte plates, and each dilution factor is inoculated 6 holes, and every hole 0.1ml is containing 5%CO
2, incubator is cultivated and is observed 3-5 day at 37 DEG C, calculates TCID according to Reed-Muench method
50.Every 1ml viral level is 10
9tCID
50.
Hog cholera venom viral level is measured: the virus liquid of results is got to 1ml physiological saline solution multiple proportions serial dilution, get 6 × 10
5doubly, 7 × 10
5doubly, 8 × 10
5doubly, 9 × 10
5doubly, 10
6doubly, 1.1 × 10
6doubly 6 dilution factors, ear vein injection body weight is two of 1.5-3kg rabbit, every 1ml.After family's rabbit inoculation, survey body temperature 1 morning and afternoon, after 48 hours, surveyed body temperature 1 time every 6 hours, carries out synthetic determination according to body temperature reaction and counteracting toxic substances result.Every 1ml viral level is 900,000 rabbit infective doses.
4. join Seedling, subpackage and lyophilizing
By the pig blue-ear disease being up to the standards, hog cholera virus liquid, mix with the volume ratio of 1: 5, be mixed in same container, add freeze drying protectant, fully shake up, quantitative separating, carries out rapidly after subpackage getting product after lyophilisation.Producing 1001,1002 and 1003,1001 batches every part reproductive and respiratory syndrome virus content of three batches of vaccines is continuously 10
6.5tCID
50, swine fever virus content is 9000 rabbit infective doses; 1002 batches every part reproductive and respiratory syndrome virus content is 10
6.2tCID
50, swine fever virus content is 9000 rabbit infective doses; 1003 batches every part reproductive and respiratory syndrome virus content is 10
6.3tCID
50, swine fever virus content is 9000 rabbit infective doses.
5. the safety of vaccine and efficacy test
Safety examination: reproductive and respiratory syndrome part, three batches of vaccines are done to suitable dilution (10 part/ml) with normal saline respectively, with 15 of 3-4 PRRS virus antigen in age in week, negative antibody piglets.Every batch of vaccine is with 5, the vaccine 1ml that each intramuscular injection has been diluted, and take temperature day by day, observes 21, does not all occur any part and the general reaction that are caused by vaccine.Meanwhile, select through the detection of neutralization test method and (before note Seedling, observe 5-7 day, each survey morning and afternoon every day body temperature 1 time without 12 of the healthy susceptible wean pigs of hog cholera antibody.Select body temperature, spirit, the normal person's use of appetite).Every batch of freeze dried vaccine sample mixed in equal amounts, the head indicating by label part is diluted to every 1ml with sterile saline and contains 6 part vaccines, 4 of intramuscular injection pigs, every 5ml (containing 30 using dosages).After note Seedling, body temperature is respectively observed and surveyed to every day at the upper and lower noon 1 time, observes 21.Body temperature, spirit, appetite do not have significant change compared with before note Seedling, and vaccine is judged to qualified.
Efficacy test: reproductive and respiratory syndrome part, with age in 4-6 week 30 of PRRS virus antigen, negative antibody pigs, 10 of the every batch of vaccines, wherein 1 part of 5 intramuscular injection vaccines, another 5 in contrast, isolated rearing under equal conditions.After 28 days, the virus-culturing fluid (>=10 of strong malicious NVDC-JXA1 strain for all pig muscle injection inspections
4.5tCID
50/ ml) 3ml, every day thermometric observing 21.5 all morbidities of contrast pig, and 3 death, 5 full guard of immune swine.Swine fever part, presses the dated head part of label for every batch, with physiological saline solution, every part vaccine is diluted to 9000 times, and ear vein injection body weight is 2 of 1.5-3kg rabbit, every 1ml.After family's rabbit inoculation, respectively survey body temperature morning and afternoon 1 time, after 48 hours, surveyed body temperature 1 time every 6 hours, carry out synthetic determination according to body temperature reaction and counteracting toxic substances result.
1. after rabbit vaccination, body temperature reaction normal is as follows:
Sizing 48～96 hours incubation periods of thermal response (++), body temperature rise is obvious curve, has at least 3 temperature time to exceed room temperature more than 1 DEG C, and delays 18～36 hours.As delay more than 42 hours necessary counteracting toxic substances, the reactionless sizing heat that is judged to after counteracting toxic substances.
Slight fever is reacted 48～96 hours (+) incubation periods, and body temperature rise is obvious curve, has at least 2 temperature time to exceed room temperature more than 0.5 DEG C, and delays 12～36 hours.
48～96 hours incubation periods of suspicious reaction (±), it is indefinite that temperature curve rises and falls, and delays less than 12 hours; Or incubation period more than 24 hours, less than 48 hours and exceed 96 hours to 120 hours and occur thermal response.
Body temperature reaction is secondary peak, and once peak meets sizing thermal response (++) or slight fever reaction (+) standard person, must counteracting toxic substances.When reactionless after counteracting toxic substances, this rabbit thermal response can be judged to sizing heat or slight fever reaction.
Reactionless (-) body temperature is normal.
2. result is judged:
After note Seedling, when 2 rabbit are all sizing thermal response (++), or 1 rabbit is when being sizing thermal response (++), another 1 rabbit and being slight fever reaction (+), and vaccine is judged to qualified.
After note Seedling, when 1 rabbit is sizing thermal response (++) or slight fever reaction (+), another 1 rabbit is suspicious reaction (±); Or 2 rabbits are while being all slight fever reaction (+), can be after note Seedling counteracting toxic substances (inoculate fresh spleen and drench poison or lyophilizing poison) on the 7th～10.When counteracting toxic substances, add 2 of contrast rabbits, counteracting toxic substances dosage is 50～100 times of Emulsions.Every rabbit ear vein injection 1ml.
Body temperature reaction normal after counteracting toxic substances is as follows:
24～72 hours incubation periods of thermal response (+), body temperature rise is obvious curve, exceedes room temperature more than 1 DEG C, delays 12～36 hours.
Suspicious reaction (±) incubation period, it is indefinite that temperature curve rises and falls less than more than 24 hours or 72 hours, delays less than 12 hours or exceed 36 hours and do not decline.
Reactionless (-) body temperature is normal.
After counteracting toxic substances, when 2 contrast rabbits are all sizing thermal response (++), or 1 rabbit is sizing thermal response (++), and another 1 rabbit is slight fever reaction (+), and 2 note all reactionless (-) of Seedling rabbit, vaccine is sentenced qualified.
After note Seedling, be sizing heat (++) or slight fever reaction (+) if any 1 rabbit, another 1 rabbit is suspicious reaction (±) or without thermal response (-), can adopt and cut open the method for killing or adopting painstaking effort isolated viral suspicious reaction rabbit or reactionless rabbit, distinguish whether inapparent infection; Or after note Seedling, 2 rabbits are all slight fever reaction, also can be to 1 rabbit isolated viral wherein.Method is: after vaccination, between 96～120 hours, rabbit is cutd open and killed, take spleen, make 50 times with normal saline and dilute Emulsions (spleen Emulsion should be aseptic), or take painstaking effort (whole blood), inoculate 2 rabbit, every rabbit ear vein injection 1ml.All have 1 rabbit to occur sizing thermal response (++) 24～72 hours incubation periods, and it is qualified that vaccine can be judged to.As a result after three batches of vaccine injections, every batch of 2 rabbit all present sizing thermal response (++), or 1 rabbit is when being sizing thermal response (++), another 1 rabbit and being slight fever reaction (+), and vaccine is judged to qualified.
6. immunoprotection test
Head part that three batches of vaccines are indicated by label with normal saline is respectively diluted to every 1ml containing 1 part vaccine, select through neutralization test method and detect without hog cholera antibody, and 60 of the healthy susceptible wean pigs of PRRS virus antigen, negative antibody, 20 of the every batch of vaccines, wherein 1 part of 10 intramuscular injection vaccines, another 10 in contrast, isolated rearing under equal conditions.After 14 days, wherein 5 note Seedling pigs and 5 contrast pigs, injection swine fever crossdrift is blood poison 1ml (10
5minimum lethal dose, purchased from China Veterinery Drug Inspection Office, deposit number CVCC AV1411), observe 16.Contrast pig all falls ill, and has 4 death at least, and immune swine is strong living all.Every batch of other 5 the note Seedling pig after note Seedling 28 days, the virus-culturing fluid (>=10 together with the inspection of 5 contrast pig muscle injections of residue with strong malicious NVDC-JXA1 strain (be disclosed in CN 101045917A, preserving number is CGMCCNO.1964)
4.5tCID
50/ ml) 3ml, every day thermometric observing 21.5 all morbidities of contrast pig, and at least 4 death, 5 full guard of immune swine.Concrete outcome is in table 1.
The test of table 1 bigeminy Seedling immunoprotection
Conclusion (of pressure testing): inject bigeminal live vaccine of the present invention, 1 part dilution 1ml of a shot, be equivalent to high-pathogenicity porcine reproductive and respiration syndrome and swine fever and inject respectively the effect of 1 part 2ml use in conjunction, save immune drug use amount, lower the Animal stress reaction that multiple injection causes, reduced cost.Meanwhile, for mixed infection reproductive and respiratory syndrome and swine fever pig, when two kinds of single Seedlings immunity, need interval about 1 week, to reduce the impact of immunostimulation on immune effect! The present invention's one pin is prevented two diseases, has reduced immune time, has reduced immunostimulation, and immune effect is improved.
6. bigeminy Seedling is with the immune effect comparative test of single Seedling
1) preparation of blue-ear disease vaccine
Get well-grown Marc-145 cell (purchased from China Veterinery Drug Inspection Office), with trypsinization, and be inoculated in cell bottle, the strain of PRRS virus JXA1-R seed culture of viruses is done to 10 times of dilutions by MEM (production of the Invitrogen company) maintenance medium containing 1%v/v hyclone (production of PAA company) simultaneously, by 5% volume inoculum concentration virus inoculation, 37 DEG C of cultivations, observation of cell pathological changes every day (CPE), in the time that appearring in 70% above cell, receives CPE poison, freeze thawing 3 times, results virus.10 times of serial dilutions of MEM culture medium by the virus liquid of results with serum-free, get 10
-6, 10
-7, 10
-8, 10
-9, 10
-10, 10
-116 dilution factors, are inoculated in respectively 96 porocyte plates, and each dilution factor is inoculated 6 holes, and every hole 0.1ml is containing 5%CO
2, incubator is cultivated and is observed 3-5 day at 37 DEG C, calculates TCID according to Reed-Muench method
50.Virus titer is that every 1ml viral level is 10
9tCID
50.
The virus liquid being up to the standards is added to freeze drying protectant, fully shake up, quantitative separating, carries out rapidly after subpackage getting product after lyophilisation.Produce continuously three crowdes of vaccine L001, L002 and L003.
2) preparation of swine Fever Vaccine
With α-MEM (production of the Invitrogen company) cell maintenance medium containing 1%v/v hyclone (production of PAA company), be the viral suspension that fever virus lapinized Chinese Strain seed culture of viruses is made 0.3%v/v by swine fever virus, inoculate well-grown ST cell (purchased from Chinese Typical Representative culture collection center) monolayer, put 37 DEG C and continue to cultivate.After connecing poison, within 3rd, do to gather in the crops and change liquid for the first time, change liquid once later every results on the 2nd, gather in the crops 3 times, the virus liquid of results is placed in-20 DEG C of following preservations.The virus liquid of results is got to 1ml physiological saline solution multiple proportions serial dilution, get 6 × 10
5doubly, 7 × 10
5doubly, 8 × 10
5doubly, 9 × 10
5doubly, 10
6doubly, 1.1 × 10
6doubly 6 dilution factors,, ear vein injection body weight is two of 1.5-3kg rabbit, every 1ml.After family's rabbit inoculation, survey body temperature 1 morning and afternoon, after 48 hours, surveyed body temperature 1 time every 6 hours, carries out synthetic determination according to body temperature reaction and counteracting toxic substances result.The every 1ml of cell venom is containing 900,000 rabbit infective doses of virus.
The virus liquid being up to the standards is added to freeze drying protectant, fully shake up, quantitative separating, carries out rapidly after subpackage getting product after lyophilisation.Produce continuously three crowdes of vaccine W001, W002 and W003.
3) bigeminy Seedling is with the immune effect comparative test of single Seedling
Head part that bigeminy vaccine 1001,1002 and 1003 in three crowdes of embodiment 1 is indicated by label with normal saline is respectively diluted to every 1ml containing 1 part vaccine, select through neutralization test method and detect without hog cholera antibody, and 60 of the healthy susceptible wean pigs of PRRS virus antigen, negative antibody, 20 of the every batch of vaccines, wherein 1 part of 10 intramuscular injection vaccines, another 10 in contrast, isolated rearing under equal conditions.After 14 days, wherein 5 note Seedling pigs and 5 contrast pigs, injection swine fever crossdrift is blood poison 1ml (10
5minimum lethal dose), observe 16.Contrast pig all falls ill, and has 4 death at least, and immune swine is strong living all.Every batch of other 5 the note Seedling pig after note Seedling 28 days, the virus-culturing fluids (>=10 together with the inspection of 5 contrast pig muscle injections of residue with strong malicious NVDC-JXA1 strains
4.5tCID
50/ ml) 3ml, every day thermometric observing 21.5 all morbidities of contrast pig, and at least 4 death, 5 full guard of immune swine.
Three crowdes of pig blue-ear disease vaccine L001, L002 and L003 are diluted to every 1ml containing 1 part vaccine with normal saline by the dated head part of label respectively, select 30 of 4-6 pig blue-ear disease antigen in age in week, negative antibody pigs, 10 every batch, wherein 1 part of 5 intramuscular injection vaccines, another 5 in contrast, isolated rearing under equal conditions.After 28 days, the virus-culturing fluid (>=10 of strong malicious NVDC-JXA1 strain for all pig muscle injection inspections
4.5tCID
50/ ml) 3ml, every day thermometric observing 21.5 all morbidities of contrast pig, and at least 2 death, 5 at least 4 head protections of immune swine.
Three crowdes of swine Fever Vaccine W001, W002 and W003 are pressed to 3000 times of part of every part dilutions of head that label indicates with normal saline respectively, select without 15 of the healthy susceptible pigs of swine fever neutralizing antibody, 5 every crowd, 2 of intramuscular injection, every 1ml.After 14 days, together with 3 of contrast pigs, injection swine fever crossdrift is blood poison 1ml (10
5minimum lethal dose), observe 16.Contrast pig all falls ill, and at least dead 2, immune swine is strong living or slightly body temperature reaction all, but be qualified without swine fever clinical symptoms.
Table 2 bigeminy Seedling is with single Seedling immune effect comparative test result
Therefore, prove by above embodiment, bigeminy vaccine of the present invention, first passage makes two kinds of antigens reach appropriate proportioning, and two kinds of antigens are not disturbed mutually, and vaccine potency at least reaches or exceeds the effect of original single Seedling.
The foregoing is only preferred embodiment of the present invention, in order to limit the present invention, within the spirit and principles in the present invention not all, any amendment of doing, be equal to replacement, improvement etc., within all should being included in protection scope of the present invention.
Claims (3)
1. a bigeminy vaccine for prevention or treatment pig blue-ear disease and swine fever, is characterized in that, comprises following composition: pig blue-ear disease antigenic component, swine fever antigen composition and vaccine freeze-drying protective agent, wherein said pig blue-ear disease antigenic content is 10
6.2tCID
50/ head part～10
6.5tCID
50/ head part, described swine fever antigen content is 9000 rabbit infective doses/head part; Described pig blue-ear disease antigen is PRRS virus JXA1-R strain, and described swine fever antigen is fever virus lapinized Chinese Strain.
2. a preparation method for bigeminy vaccine as claimed in claim 1, is characterized in that, comprises the following steps: use passage cell to cultivate PRRS virus and swine fever virus, to obtain pig blue-ear disease antigen and swine fever antigen; Be mixed in proportion two kinds of antigenic components, be aided with freeze drying protectant and be prepared into freeze dried vaccine; Wherein, described pig blue-ear disease antigenic content is 10
6.2tCID
50/ head part～10
6.5tCID
50/ head part, described swine fever antigen content is 9000 rabbit infective doses/head part; Described pig blue-ear disease antigen is PRRS virus JXA1-R strain, and described swine fever antigen is fever virus lapinized Chinese Strain.
3. a bigeminy vaccine as claimed in claim 1 purposes in the medicine of preparation prevention or treatment pig blue-ear disease and swine fever.
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| CN102727883A (en) * | 2011-05-27 | 2012-10-17 | 华威特（北京）生物科技有限公司 | Combined live vaccine against porcine reproductive and respiratory syndrome and swine fever, and application thereof |
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| CN102727883A (en) * | 2011-05-27 | 2012-10-17 | 华威特（北京）生物科技有限公司 | Combined live vaccine against porcine reproductive and respiratory syndrome and swine fever, and application thereof |
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