Preparation method of rabies inactivation vaccine for livestock and stabilizing agent contained in vaccine
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A technology of rabies vaccine and inactivated vaccine, which is applied in the field of biological products, can solve the problems of low antigen purity, difficulty in balancing stability and quality, and achieve the effect of improving antigen purity, facilitating large-scale and efficient production, and reducing volume
Active Publication Date: 2019-10-18
WUHAN KEQIAN BIOLOGY CO LTD
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[0005] The present invention provides a preparation method of veterinary rabies inactivated vaccine and the stabilizer contained in the vaccine, by optimizing the concentration and purification operations in the production process, and using The specific stabilizer overcomes the defects of low purity of rabies vaccine antigen in the prior art, and difficulty in balancing stability and quality
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Embodiment 1
[0057] This embodiment provides a method for preparing an inactivated rabies vaccine for animals. The specific steps are as follows:
[0058] 1. Virus preparation process: Take BHK-21 cells in pure suspension culture at 37°C, inoculate with 0.05 MOI of rabies virus, harvest 220L of virus liquid, use continuous flow centrifugation, rotate speed 8000rpm, and set the sample flow rate to 150L / h. Then use multi-stage microfiltration membrane to filter: first filter with 8um filter membrane made of cellulose acetate, and then filter with 3um+0.8um composite filter membrane to obtain 200L of virus supernatant.
[0059] 2. Virus concentration process:
[0060] (1) Put the virus supernatant in the antigen treatment tank, and circulate the virus supernatant in the antigen treatment tank at 55L / min for 2min, adjust the pump flow rate to open the permeate end, and control the permeate end to flow at 0.8L / min Continue to permeate until the volume of the virus supernatant is concentrated to 5L to...
Embodiment 2
[0073] This embodiment provides a method for preparing a rabies vaccine for veterinary use. The specific steps are the same as those in Example 1, except that step (2) of the virus concentration process is adjusted to:
[0074] Add 50L PBS solution to the crude concentrate, set the pump flow rate to 45L / min, circulate for 10 minutes, adjust the pump speed to open the permeate end, control the permeate end to continuously permeate at a flow rate of 1.8L / min, and concentrate to 20L in volume When, reduce the pump speed to adjust the flow rate of the permeate end to 0.8L / min, continue to permeate until the volume is concentrated to 5L, close the permeate end; this step is repeated 3 times to obtain the virus concentrate.
Embodiment 3
[0076] This example provides a method for preparing a rabies vaccine for veterinary use. The specific steps are the same as in Example 1, except that the stabilizer is hydrolyzed milk protein 30mg / ml, lysine 60mg / ml, and proline tert-butyl ester 25mg / ml And sorbitol 25mg / ml.
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Abstract
The invention provides a preparation method of a rabies inactivation vaccine for livestock and a stabilizing agent contained in the vaccine. According to the preparation method, a virus concentrationprocess is that (1) a virus supernate continuously penetrates through a 300 KD membrane bag at the flow speed of 0.8-1 L/min, the virus is concentrated until the volume of the virus is 1/50-1/40 of the original volume, and coarse concentration liquid is obtained; (2) a PBS solution with the volume 10-15 times larger than that of the coarse concentration liquid is added into the coarse concentration liquid, the mixture penetrates through the 300 KD membrane bag at the flow speed of 1.8-2 L/min under the concentration pressure of 0.5-1.0 bar, under the condition that the mixture is concentrateduntil the volume of the mixture is 1/4-2/5 of the original volume of the mixture before penetration, the flow speed is adjusted to 0.8-1 L/min, and continuous concentration is conducted until the volume of the mixture is the same as the original volume of the coarse concentration liquid. According to the preparation method, by optimizing the virus concentration process, different membrane-penetration flow speeds are adopted for different antigen concentrations, in the concentration process, 30-50% of impure protein can be removed, the virus titer is kept at 90% or above, and the prepared vaccine is more stable and more efficient.
Description
Technical field [0001] The invention relates to the field of biological products, in particular to a preparation method of an inactivated rabies vaccine for animals and a stabilizer contained in the vaccine. Background technique [0002] Rabies (Rabies) is a zoonotic infection caused by the rabies virus (Rabiesvirus) of the Rhabdoviridae family and the genus Rabies virus. It is also known as waterphobia, rabies, etc. It is a kind of damage to the central nervous system. Acute viral infectious disease, all warm-blooded animals, including humans, can be infected. The disease is mainly infected by rabies virus liquid carried in animal saliva invading the body. Dogs are the main storage host and transmission vector of rabies virus in my country . The incubation period of rabies is generally from half a month to three months, most of which are concentrated in 30 to 90 days. Once rabies occurs, its development speed is very fast, mostly in 3 to 5 days, rarely more than 10 days, the case...
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