CN1958068A - Immunoglobulin of anti canine distemper in use for dogs, fox, and ermine - Google Patents
Immunoglobulin of anti canine distemper in use for dogs, fox, and ermine Download PDFInfo
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- CN1958068A CN1958068A CN 200610131635 CN200610131635A CN1958068A CN 1958068 A CN1958068 A CN 1958068A CN 200610131635 CN200610131635 CN 200610131635 CN 200610131635 A CN200610131635 A CN 200610131635A CN 1958068 A CN1958068 A CN 1958068A
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- canine distemper
- ermine
- dog
- immunoglobulin
- vulpes
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Abstract
An immunoglobulin for preventing canine distemper of dog, fox and marten is prepared from the low virulent strain Rockborn of cinine distemper through cell culture, concentrating and proportionally mixing it with complete Frennd's adjuvant. It features that the glycoside-peptide injection is used as its immunopotentiator.
Description
Technical field
The invention belongs to biological technical field, be specifically related to the immunoglobulin of a kind of treatment and prevention dog, Vulpes, ermine canine distemper, international Patent classificating number is:: A61K39/395.
Background technology
Canine distemper is a kind of acute, infectious disease that infectiousness is extremely strong.Canidae (dog, Vulpes etc.) and Mustelidae (ermine etc.) all infect.To dog, mainly invade pup, seldom see adult dogs, break out canine distemper first in the area that does not have canine distemper, mortality rate can reach 90%; To mink, canine distemper claims the ermine pestilence, when ermine belongs to animal and is the source of infection, can cause wide-scale distribution in the ermine group through 3～4 weeks, serious symptom, and the mortality rate height, sometimes up to 80%～90%, joey particularly.Therefore, canine distemper is to support dog industry, the maximum infectious disease of economic animal aquaculture harm.
Dog, Vulpes, ermine are to the canine distemper virus sensitivity.Can produce strong persistent immunity after the canine distemper rehabilitation.To its prophylactic immunization, can produce anti-canine distemper specific antibody, claim immunoglobulin again.Specific reaction can take place with the canine distemper antigenic determinant that stimulates them to produce in antibody.Different types of animal is the fact that people are familiar with already to the different resistances and the sensitivity of same virus, and equus (horse, donkey, mule) is to canine distemper virus susceptible not, but under immunogenic stimulation, presents the immunoreation of high special equally.With equus prepare dog, Vulpes, not only volume of production is big with immunoglobulin for ermine, and can avoid other viral infectious of dog, Vulpes, ermine to cause artificial infection by preparation process.
Give dog, Vulpes, the passive input immunoglobulin of ermine, often can make its infection present prevention and unique therapeutical effect canine distemper.But the immunoglobulin that must make input reaches the required level of protection.This passive immunity is of short duration, is no more than for 2 weeks, and therefore being used for urgent prevention is that reasonably it is certain being used for the early treatment, also is unique.
The canine distemper vaccine inoculation of dog, Vulpes, ermine is still the most reasonably measure of prevention canine distemper.
The canine distemper inactivated vaccine, the antibody titer of producing for dog, Vulpes, ermine immunity descends very fast, and is similar to colostral antibody, descended 50% in per 10 days, no matter whether inactivated vaccine adds adjuvant, the antibody duration, inactivated vaccine was seldom used generally speaking all than attenuated vaccine much shorter.
Canine distemper live vaccine is particularly used the weak malicious Seedling of cell culture of Vero cells produce, and inoculation back antibody titer is very high, and immunity is strong and lasting.The main problem that exists is: because of fearing the maternal antibody influence, can not immunity inoculation to the young beast before weaning, as be strictly that maternal antibody protecting on the desired level, immunity inoculation necessarily causes immuning failure.If but because of just the Ruzhong maternal antibody is low, young beast does not reach the protection desired level by the maternal antibody that colostrum obtains, the so young beastly very big possibility that infects canine distemper virus that just has.And for example after vaccination, need could produce immunity through 2～3 weeks, about inoculation 4 weeks of back, antibody titer peaks.Therefore in 3～4 weeks after the vaccination, young beast just has the very big possibility that infects canine distemper virus; Canine distemper attenuated for another example minimum immunity amount is 10
4TCID
50, using dosage is that 20 minimum immune dosages are 2 * 10
5TCID
50Since canine distemper attenuated responsive to visible light and temperature etc., often make the poison amount of immunity inoculation make immuning failure inadequately.In addition after first immunisation, should go secondary even three immunity at interval in 21 days again, immune time is not enough, so that in the immunity later stage of effect duration, the condition of generation canine distemper has been arranged.
Dog, Vulpes, ermine having been sent out canine distemper, promptly inoculated with canine distemper live vaccine, promptly inoculated with the measles attenuated vaccine, is not very big to the prevention of healthy dogs, Vulpes, ermine on every side with to the therapeutical effect of morbidity dog, Vulpes, ermine.Because canine distemper live vaccine is promptly inoculated, under the situation of dog, Vulpes, ermine health, 2～3 weeks could produce immunity, before 2～3 weeks, fell ill in the epidemic-stricken area and did not fall ill, and the danger of being annihilated is all arranged.
Summary of the invention
Purpose of the present invention just provides a kind ofly has prevention and a kind of dog of therapeutical effect, Vulpes, ermine with anti-canine distemper immunoglobulin to canine distemper.
Immunogen of the present invention adopts the canine distemper attenuated Rockborn strain of introducing from Russia, through the rejuvenation of dog peritoneal macrophage, uses the Vero cell proliferation, and every 0.1ml is toxic 〉=and 10
5TCID
50, through the freezing ultra-lowing centrifuge differential centrifugation concentrate every 0.1ml toxic 〉=10
7.0TCID
50Be mixed into antigen with Freund's complete adjuvant equivalent (1: 1);
The present invention adopts the Jilin whale to resemble the Heterosides peptide injection of animal pharmaceutcal corporation, Ltd development, and (denomination of invention is: a kind of Heterosides peptide injection that improves the animal immune function, number of patent application is: 2,005 1 0016626.2) be immunostimulant, 5ml/ props up.
The present invention selects horse, donkey, mule immune animal.3～8 years old, build is big, disposition is docile, person in good health; Equine infectious anemia, glanders quarantine negative patient; Observe healthy person through isolated rearing before the immunity.
Immune programme for children
Horses are implemented fundamental immunity and reinforced immunological.
1, fundamental immunity immunity for the first time is a fundamental immunity, antigen 1,4ml, 4 intradermal injections in both sides, back, Heterosides peptide injection 5.0ml, intragluteal injection.
2, after the reinforced immunological fundamental immunity 21 days, 2 reinforced immunologicals of row, each 7 days at interval, the amount of immunizing antigen 1 was 8ml for the first time, 4 intradermal injections in both sides, back, every some 1ml, every some 2ml of shoulder both sides lymph node; Immunity for the second time, canine distemper attenuated cell culture concentrating virus liquid 3ml, vein is slowly injected.During each reinforced immunological, intragluteal injection Heterosides peptide injection 5.0ml.
Blood count and blood-letting standard immune the last time back 14 days, are surveyed the anti-canine distemper NAT in immune horse, donkey, the mule serum with fixed virus dilute serum method.Treat that NAT reaches at 1: 1024 o'clock, can take a blood sample.
Blood sampling and serum separate horse, donkey, the mule that meets the blood-letting standard, and blood sampling connects 15～18ml/kg body weight for the first time, and 3～5 light carry out the blood sampling second time at interval, and the secondary blood sampling volume lacks 300～400ml than a blood sampling volume; After the secondary blood sampling, immune animal carried out the second journey reinforced immunological after having a rest 20 days, and the rest may be inferred.
The blood of gathering pressurization nature coagulation separation of serum.
Making with extra care of serum
Serum → doubly measure adding distil water, transfer PH to 3.4～3.6 with 2mol/L hydrochloric acid, amount by every ml6～9 units adds pepsin, add methylbenzyl by total amount 0.2%, 29～31 ℃ of digestion 2～21 hours → add ammonium sulfate by 15% of total amount, the sodium hydroxide of 2mol/L is transferred PH4.5～4.7,56～58 ℃ of water-baths 30 minutes, be cooled to below 43 ℃ and filter, get filtrate → filtrate and transfer PH to 7.1～7.3 with the 2mol/L caustic lye of soda, add ammonium sulfate by total amount 20%, canvas filters, get precipitation → precipitation and add 5～6 times of dissolved in distilled water, add 10% alum solution by total amount, making Alumen content is 1%, with the caustic lye of soda of 2mol/L, transfers PH7.7～7.8, leave standstill and made it precipitation in 2～4 hours, canvas filters, and the filtrate of getting filtrate → collection adds ammonium sulfate by 38% of total amount, and canvas filters, abandon supernatant, precipitation press dry uses the bag filter bag distribution packaging, and every bag of about 200～250g adds a little chloroform, is placed on the interior serum of 48～72 hours → collecting bag in the gutter channel, add chloroform by 0.5% of total amount, 0.85% adds sodium chloride and 0.0035% adds thimerosal, shakes up filtration sterilization.Be the semi-finished product of refining anti-canine distemper immunoglobulin.
Semi-finished product titration and packing are surveyed half-finished anti-canine distemper NAT (log with fixed virus dilute serum method
10SN
50) SN 〉=2048; The regulation of product SN 〉=1300 of tiring, additional 20%, therefore semi-finished product are diluted to SN 〉=1600, quantitatively packing.
The relation of the strong malicious counteracting toxic substances protection of dog, Vulpes, the clear anti-canine distemper NAT of Ermine blood and canine distemper; Dog, Vulpes, the clear SN of Ermine blood 〉=protection more than 50,90%, the above protection in SN28～40,50%;＜16 do not protect.
Use product of the present invention, press the 1ml/kg body weight and give dog, Vulpes, ermine injection, behind the 4h, the anti-canine distemper antibody of its serum SN 〉=50, sustainable more than 14 days.Therefore, the prevention using dosage of dog, Vulpes, ermine is the 1ml/kg body weight, and dog, Vulpes, ermine to sending out canine distemper, the early treatment is the 2ml/kg body weight, state of an illness weight person or biphasic injection or increasing injected dose.
In the plant of dog, Vulpes, ermine, when the canine distemper epidemic situation takes place, urgent protective inoculation, protection more than 96%; The isolation early treatment of morbidity cures more than 90%; Outpatient service morbidity dog cooperates auxiliary treatment, 2～3 treatments of heavy person, cure rate about 85%.
Good effect of the present invention
1, safety is produced in the process for refining of immunoglobulin well, uses pepsin digestion serum; Transfer PH4.5～4.7,56～58 ℃ of water-baths 30 minutes, can eliminate foreign sera to dog, Vulpes, the possible anaphylaxis (anaphylaxis is rare) of ermine, can fundamentally guarantee security of products effective deactivation (the exogenesis viral pollution verifies as feminine gender) that dog, Vulpes, the responsive virus of ermine may be polluted.With joey muscle or the subcutaneous injection of 5 times of using dosages (5ml/kg), observe all having no adverse reaction in two weeks to dog, Vulpes, ermine.
2, (every 0.1ml toxic 10 with canine distemper attenuated for anti-canine distemper NAT height
5TCID
50), concentrate 100 times through the differential centrifugation method, add adjuvant (Fu Shi Freund's complete adjuvant), cooperate immunostimulant (Heterosides peptide injection), press optimal path (Intradermal, lymph node, vein) and dosage, through 1 basis immunity and twice reinforced immunological, and make the anti-canine distemper NAT of serum 〉=1024 of horse, donkey, mule, the approaching level for preparing immunoglobulin with dog as immune animal.The anti-canine distemper NAT of product 〉=2048 in the middle of process for refining is guaranteed again.Better solved with horse and prepared the anti-canine distemper immunoglobulin not high problem of tiring.
3, anti-canine distemper NAT in clear of the accuracy dog of using dosage, Vulpes, Ermine blood 〉=50 o'clock use the canine distemper strong virus attack, protects more than 90%; The standard of the anti-canine distemper neutralizing antibody of product is 〉=1024; Product with the 1ml/kg body weight is given dog, Vulpes, ermine intramuscular injection, behind the 4h, and the anti-canine distemper NAT of serum 〉=50.And continue more than 14 days.
4, the foster dog field (3,300 of dogs) of canine distemper takes place in clinical trial result fortunately, minkery (5,30,000 of ermines) fox farm (1,600 of foxes), and to the early stage cure rate 90% of isolation of morbidity, all the other all urgent prophylactic immunizations obtain protection more than 96%.Toy hospital admission treatment canine distemper disease dog, owing to do not reach the early treatment, so after auxiliary treatment and secondary (have three times) injection, cure rate is about 85%.Anti-in a word canine distemper immunoglobulin is specific to the urgent prevention and the treatment of canine distemper.
5, preservation convenient transportation this product is the immunoglobulin with the serum extraction of immune horse, donkey, mule, be colourless transparent liquid, under 4 ℃ of conditions, it is 1 year that anti-canine distemper NAT SN reduces to time of 1300 from 1600, promptly 4 ℃ of storage lives are 1 year, transport also more convenient.
The specific embodiment
Immunogen of the present invention is to adopt the canine distemper attenuated Rockborn strain of introducing from Russia, through the rejuvenation of dog peritoneal macrophage, uses the Vero cell proliferation, every 0.1ml is toxic 〉=and 10
5TCID
50, through the freezing ultra-lowing centrifuge differential centrifugation concentrate every 0.1ml toxic 〉=10
7.0TCID
50Be mixed into antigen with the complete Freund's complete adjuvant equivalent of Fu Shi (1: 1);
The Heterosides peptide injection that the present invention adopts the Jilin whale to resemble the development of animal pharmaceutcal corporation, Ltd is an immunostimulant, and 5ml/ props up.
Immune programme for children
The present invention selects horse, donkey, mule immune animal.3～8 years old, build is big, disposition is docile, person in good health; Equine infectious anemia, glanders quarantine negative patient; Observe healthy person through isolated rearing before the immunity.
Horses are implemented fundamental immunity and reinforced immunological.
1, fundamental immunity immunity for the first time is a fundamental immunity, antigen 1,4ml, 4 intradermal injections in both sides, back, Heterosides peptide injection 5.0ml, intragluteal injection.
2, after the reinforced immunological fundamental immunity 21 days, 2 reinforced immunologicals of row, each 7 days at interval, the amount of immunizing antigen 1 was 8ml for the first time, 4 intradermal injections in both sides, back, every some 1ml, every some 2ml of shoulder both sides lymph node; Immunity for the second time, canine distemper attenuated cell culture concentrating virus liquid 3ml, vein is slowly injected.During each reinforced immunological, intragluteal injection Heterosides peptide injection 5.0ml.
Blood count and blood-letting standard immune the last time back 14 days, are surveyed the anti-canine distemper NAT in immune horse, donkey, the mule serum with fixed virus dilute serum method.Treat that NAT reaches at 1: 1024 o'clock, can take a blood sample.
Blood sampling and serum separate horse, donkey, the mule that meets the blood-letting standard, and blood sampling connects 15～18ml/kg body weight for the first time, and 3～5 light carry out the blood sampling second time at interval, and the secondary blood sampling volume lacks 300～400ml than a blood sampling volume; After the secondary blood sampling, immune animal carried out the second journey reinforced immunological after having a rest 20 days, and the rest may be inferred.
The blood of gathering pressurization nature coagulation separation of serum.
Making with extra care of serum
Serum → doubly measure adding distil water, transfer PH to 3.4～3.6 with 2mol/L hydrochloric acid, amount by every ml6～9 units adds pepsin, add methylbenzyl by total amount 0.2%, 29～31 ℃ of digestion 2～21 hours → add ammonium sulfate by 15% of total amount, the sodium hydroxide of 2mol/L is transferred PH4.5～4.7,56～58 ℃ of water-baths 30 minutes, be cooled to below 43 ℃ and filter, get filtrate → filtrate and transfer PH to 7.1～7.3 with the 2mol/L caustic lye of soda, add ammonium sulfate by total amount 20%, canvas filters, get precipitation → precipitation and add 5～6 times of dissolved in distilled water, add 10% alum solution by total amount, making Alumen content is 1%, with the caustic lye of soda of 2mol/L, transfers PH7.7～7.8, leave standstill and made it precipitation in 2～4 hours, canvas filters, and the filtrate of getting filtrate → collection adds ammonium sulfate by 38% of total amount, and canvas filters, abandon supernatant, precipitation press dry uses the bag filter bag distribution packaging, and every bag of about 200～250g adds a little chloroform, is placed on the interior serum of 48～72 hours → collecting bag in the gutter channel, add chloroform by 0.5% of total amount, 0.85% adds sodium chloride and 0.0035% adds thimerosal, shakes up filtration sterilization.Be the semi-finished product of refining anti-canine distemper immunoglobulin.
Semi-finished product titration and packing are surveyed half-finished anti-canine distemper NAT (log with fixed virus dilute serum method
10SN
50) SN 〉=2048; The regulation of product SN 〉=1300 of tiring, additional 20%, therefore semi-finished product are diluted to SN 〉=1600, quantitatively packing.
Claims (3)
1, a kind of dog, Vulpes, ermine is characterized in that with anti-canine distemper immunoglobulin:
Immunogen is to adopt the canine distemper attenuated Rockborn strain of introducing from Russia;
Immunological adjuvant adopts Freund's complete adjuvant;
The Heterosides peptide injection that adopts the Jilin whale to resemble the development of animal pharmaceutcal corporation, Ltd is an immunostimulant.
2, a kind of dog according to claim 1, Vulpes, ermine is characterized in that with anti-canine distemper immunoglobulin:
Canine distemper attenuated Rockborn strain of immunogen and the blended volume ratio of immunological adjuvant Freund's complete adjuvant are 1: 1;
Described immunostimulant Heterosides peptide injection, 5ml/ props up.
3, a kind of dog according to claim 1, Vulpes, ermine is characterized in that with anti-canine distemper immunoglobulin:
Canine distemper attenuated Rockborn strain through the rejuvenation of dog peritoneal macrophage, is used the Vero cell proliferation, and every 0.1ml is toxic 〉=and 10
5TCID
50, through the freezing ultra-lowing centrifuge differential centrifugation concentrate every 0.1ml toxic 〉=10
7.0TCID
50
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 200610131635 CN1958068A (en) | 2006-11-15 | 2006-11-15 | Immunoglobulin of anti canine distemper in use for dogs, fox, and ermine |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 200610131635 CN1958068A (en) | 2006-11-15 | 2006-11-15 | Immunoglobulin of anti canine distemper in use for dogs, fox, and ermine |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN1958068A true CN1958068A (en) | 2007-05-09 |
Family
ID=38070021
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN 200610131635 Pending CN1958068A (en) | 2006-11-15 | 2006-11-15 | Immunoglobulin of anti canine distemper in use for dogs, fox, and ermine |
Country Status (1)
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|---|---|
| CN (1) | CN1958068A (en) |
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2006
- 2006-11-15 CN CN 200610131635 patent/CN1958068A/en active Pending
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Open date: 20070509 |