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Freeze-dried heat-resistant protective agent for porcine pseudorabies live vaccine, its preparation method and use

A technology for porcine pseudorabies and heat-resistant protective agent, applied in the field of immunity, can solve the problems of intolerance of porcine pseudorabies vaccine to freezing, heat intolerance of transportation and storage, etc., to solve the problem of low temperature cold chain, solve the problem of intolerance, Minimal loss of antigen

Active Publication Date: 2020-10-16
JIANGSU ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] Aiming at the defects of the prior art, the object of the present invention is to provide a freeze-dried heat-resistant protectant for porcine pseudorabies low-toxicity live vaccine and its use method, in order to solve the problem of porcine pseudorabies vaccine being intolerant to freezing and transporting and storing. heat resistance problem

Method used

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  • Freeze-dried heat-resistant protective agent for porcine pseudorabies live vaccine, its preparation method and use
  • Freeze-dried heat-resistant protective agent for porcine pseudorabies live vaccine, its preparation method and use
  • Freeze-dried heat-resistant protective agent for porcine pseudorabies live vaccine, its preparation method and use

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0038] Embodiment 1 Porcine pseudorabies live vaccine antigen preparation and the preparation of Hanks (containing phenol red)

[0039] Virus strain: PRV-Bartha-k61; cells: porcine testis cells (ST), all provided by the National Veterinary Biological Products Engineering Technology Research Center. Take a sample of porcine pseudorabies virus, dilute it 100 times with 2% DMEM, inoculate ST cells, culture at 37°C for 48 hours, harvest the virus liquid, freeze and thaw twice, filter with gauze, harvest the supernatant, and store the sample in Freeze at -80°C. After being tested for sterility, the virus content was determined to be 10 8.0 TCID 50 / ml, it can be used for seedling matching.

[0040] Preparation of Hanks (containing phenol red)

[0041] Hanks solution is the most commonly used inorganic salt solution and balanced salt solution (Balanced Salt Solutions, BSS) in biomedical experiments. Its preparation method is as follows:

[0042] stock solution A

[0043]

...

Embodiment 2

[0052] The mensuration of embodiment 2 heat-resistant protective agent thermal parameters

[0053] In order to better set the freeze-drying curve, the inventors measured the collapse temperature of the heat-resistant protective agent. The collapse temperature was determined by a low-temperature freeze-drying microscope, and the detection conditions were: Temp-40°C, Vac 20.0Pa, Ramp Row 3Rate 3°C / min, Limit-40°C. Specifically, it is tested by Shanghai Tofflon Co., Ltd. The test result is: protective agent -15°C.

Embodiment 3

[0054] The preparation of embodiment 3 protective agent 1 and the lyophilization of vaccine

[0055] The freeze-dried heat-resistant protective agent 1 is composed of the following components by weight percentage: 5% sucrose, 4% isomaltulose, 2% D-sorbitol, 1.5% glycerin, 3% hypromellose, 2.5% methionine, poly Poly D-lysine 0.01%, Hanks (containing phenol red), double pure water to make up to 100%.

[0056] Take the heat-resistant protective agent 1 and the qualified PRV-Bartha-k61 antigen at a volume of 1:1, mix them into 7ml vials, 2ml / bottle, and put the product half-stoppered with a butyl tripod stopper into a freeze-drying box Pre-cool the body at 4°C for 45 minutes, lower the temperature to -40°C at a rate of 1.37°C / min, and maintain the vaccine for 4-5 hours to freeze the vaccine. After vacuuming to 500mTorr, gradually increase the temperature at a rate of 1.87°C / min up to 6°C, maintain at this temperature for 2 hours, then cool down to -22°C at a rate of 2.2°C / min and...

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Abstract

The invention provides a porcine pseudorabies live vaccine freeze-drying heat-resisting protective agent, a preparation method and an application thereof. The protecting agent is characterized by being free of gelatin and proteins. The protecting agent contains the following components in percentage by weight: 2-8% of saccharose, 3-6% of isomaltulose, 1-5% of D-sorbitol, 1-5% of glycerinum, 0.1-5%of hydroxypropyl methylcellulose, 0.1-3.5% of methionine, 0.01-0.05% of poly-D-lysine, Hanks (containing phenol red) and the balance of double pure water, totaling 100%. The porcine pseudorabies livevaccine prepared by the freeze-drying heat-resisting protective agent is relatively good in heat resistance, the freeze-drying loss is 0.5-0.65Lg, the heat resistant loss at 37 DEG C for 10 days is 0.2-0.25Lg, and the storage life loss at 2-8 DEG C for 30 days is 0.7-0.9Lg. The low-temperature cold chain problem in storage and transportation of the vaccine is solved, and in particular the rural low-temperature cold chain problem in a remote region is solved.

Description

technical field [0001] The invention belongs to the technical field of immunization, and in particular relates to a freeze-dried heat-resistant protective agent for porcine pseudorabies live vaccine and its preparation method and application. Background technique [0002] Porcine pseudorabies (Pseudorabies) is caused by pseudorabies virus (Pseudorabies virus, PRV), a variety of livestock with fever, itching (except pigs), reproductive disorders, encephalomyelitis as the main symptoms of a highly contagious infectious disease. It is a useful model virus for studying herpes virus biology. The taxonomic name of pseudorabies virus is porcine herpesvirus type I, which belongs to the genus varicellaherpesvirus of the subfamily Herpesviridae. All herpesviruses have a double-stranded DNA genome, virion particles of similar size (200-250nm), similar structure (capsid, envelope, and envelope), and have a latent period in the life cycle (Roizman et al., 2001). PRVs are also used as "...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A61K9/19A61K47/26A61K47/34A61K47/38A61K47/18A61K47/10A61K39/245A61P31/22
CPCA61K9/19A61K39/12A61K47/10A61K47/183A61K47/26A61K47/34A61K47/38A61K2039/552A61P31/22C12N2710/16734
Inventor 赵艳红吕芳卢宇常晨左晓昕邓碧华张金秋
Owner JIANGSU ACAD OF AGRI SCI
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