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Preparation and freeze-dried storage method of pseudorabies standard positive serum

A technology for pseudorabies and positive serum, which is applied in the field of animal virus live vaccine inspection, can solve the problems of difficult to neutralize viruses, take a long time, and be expensive, and achieve good stability, good technical support, and long storage time.

Active Publication Date: 2014-06-18
WUHAN CHOPPER BIOLOGY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The "Regulations of the People's Republic of China on Biological Products" (2000 edition) and "The Veterinary Pharmacopoeia of the People's Republic of China" (2010 edition 3) both require the identification and testing of live pseudorabies vaccines and foreign virus testing, but due to the existing domestic pseudorabies standards Positive serum preparation technology takes a long time, low success rate, and low antibody titer, which makes it difficult to completely neutralize the corresponding virus in the vaccine identification test and exogenous virus test, making it difficult to carry out the quality inspection of the vaccine normally, and affecting the quality of the vaccine. Supervision has left hidden dangers. It is difficult and expensive to purchase foreign pseudorabies standard positive serum. In addition, there is a large demand for testing. In order to meet the needs of production and testing, it is imperative to develop a method for preparing high-potency pseudorabies standard positive serum.

Method used

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  • Preparation and freeze-dried storage method of pseudorabies standard positive serum
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  • Preparation and freeze-dried storage method of pseudorabies standard positive serum

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Experimental program
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Effect test

Embodiment 1

[0025] 1. Preparation of serum

[0026] Five Japanese big-eared white rabbits weighing 1.5-2kg (clean grade) were screened, and the antibodies to pseudorabies, rotavirus, swine fever, parvovirus, and rabbit plague were all negative by PCR and ELISA. Each rabbit’s left groin was injected subcutaneously with PRV (C strain) inactivated virus solution (10 7.0 TCID 50 / mL) 2mL, after 14 days, each rabbit’s left groin was subcutaneously boosted with PRV (C strain) inactivated virus solution (10 7.0 TCID 50 / mL) 2mL, 28 days after the first immunization, each rabbit was injected subcutaneously with PRV (C strain) virus solution (10 5.0 TCID 50 / mL) 2mL, 14 days after the third immunization, the rabbit carotid artery can be bled to collect blood and separate serum for later use. Among them, PRV (C strain) is a naturally attenuated Pseudorabies C strain, the preservation number is: CCTCC NO: V201114; preservation unit: China Center for Type Culture Collection, CHINA CENTERFOR TYPE CULTURE ...

Embodiment 2

[0038] 1. Preparation of serum

[0039] 10 Japanese big-eared white rabbits weighing 1.5-2kg (clean grade) were screened. PCR and ELISA were used to detect pseudorabies, porcine reproductive and respiratory syndrome, swine fever, porcine parvovirus disease, and rabbit fever antigen antibodies. Each rabbit’s left groin was injected subcutaneously with PRV (C strain) inactivated virus solution (10 7.0 TCID 50 / mL) 2mL, after 14 days, each rabbit’s left groin was subcutaneously boosted with PRV (C strain) inactivated virus solution (10 7.0 TCID 50 / mL) 2mL, 28 days after the first immunization, each rabbit was injected subcutaneously with PRV (C strain) virus solution (10 5.0 TCID 50 / mL) 2mL. 15 days after the third immunization, the rabbit carotid artery can be bled to collect blood and separate serum for later use.

[0040] 2. Serum neutralizing antibody titer test

[0041] The collected rabbit serum was sterilized with a 0.22 μm filter membrane and inactivated in a 56°C water bath ...

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Abstract

The invention discloses a preparation and freeze-dried storage method of pseudorabies standard positive serum. The preparation comprises the following steps: immunizing a rabbit twice with an inactivated PRV (pseudorabies virus) solution, then, immunizing the rabbit once with the inactivated PRV solution, and collecting the serum; adding a protective agent sodium chloride, trehalose, monosodium glutamate and L-arginine into the serum, fully dissolving, filtering to remove bacteria, and subpackaging; adding the subpackaged serum into a freeze dryer box, and performing freeze-drying according to a freeze-drying curve shown as fig.1 described in the specification to obtain the pseudorabies standard positive serum. For the pseudorabies standard positive serum obtained by the method disclosed by the invention, PRV neutralizing antibody titer is more than 1:512, the antibody titer remains unchanged after the pseudorabies standard positive serum is stored for 60 months at 2-8 DEG C, so that the pseudorabies standard positive serum has good stability, can fully neutralize pseudorabies virus in vaccine exogenous virus and specificity detection, and provides a good technical support for quality inspection of pseudorabies virus vaccines.

Description

Technical field [0001] The invention relates to the field of animal virus live vaccine testing, in particular to the preparation of pseudorabies standard positive serum and a freeze-drying storage method. Background technique [0002] Porcine pseudorabies (PR) is an acute infectious disease caused by Pseudorabies Virus (PRV). Its main clinical features are recessive infection in adult pigs, abortion, stillbirth and respiratory symptoms in pregnant sows. Young piglets often develop fever and diarrhea after infection, accompanied by obvious neurological symptoms and difficulty breathing. The disease spreads quickly in pigs and piglet mortality is high. It is one of the major infectious diseases faced by pig farms. my country is a country where pseudorabies is widespread. At present, many provinces (cities, districts) have reported the occurrence of this disease, which has caused huge economic losses. [0003] Vaccines play an important role in the prevention and control of porcine ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K16/18C07K16/06G01N33/96G01N33/569
Inventor 郑良益舒银辉漆世华杨思谊祝春花李玉萍谢红玲温文生李淑钰郝敬侃饶清宜王桂桂王治江
Owner WUHAN CHOPPER BIOLOGY
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