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Method for testing efficacy of porcine circovirus type 2 inactivated vaccine

A technology of vaccine and effectiveness, which is applied in the field of testing the effectiveness of porcine circovirus type 2 inactivated vaccines, can solve the problem of not truly reflecting the immune effect of porcine circovirus type 2 inactivated vaccines, large differences between batches of test results, and problems in testing Pig screening time-consuming and other issues, to achieve the effect of short time-consuming, high stability, simple operation

Pending Publication Date: 2022-07-22
成都史纪生物制药有限公司
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  • Description
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AI Technical Summary

Problems solved by technology

This method overcomes the time-consuming and labor-intensive screening of test pigs in the traditional piglet challenge method and the large differences in test results between batches. However, this method detects the antibody level, not the neutralizing antibody level, and cannot truly reflect the level of piglets Immune Effect of Circovirus Type 2 Inactivated Vaccine
[0006] Therefore, there is an urgent need to develop a method that can not only overcome the problems of time-consuming, laborious and large differences between batches of test pigs in the traditional piglet challenge method, but also more truly reflect the efficacy of porcine circovirus type 2 inactivated vaccines. Immunoefficacy Method to Test the Efficacy of PCV2 Vaccines

Method used

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  • Method for testing efficacy of porcine circovirus type 2 inactivated vaccine
  • Method for testing efficacy of porcine circovirus type 2 inactivated vaccine
  • Method for testing efficacy of porcine circovirus type 2 inactivated vaccine

Examples

Experimental program
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Effect test

preparation example Construction

[0034] The preparation method of porcine circovirus type 2 cytotoxicity is as follows:

[0035] Select the well-grown Pk15 cell monolayer, pour off the cell fluid, add the virus seed according to the inoculation amount of 5% (V / V), add the virus culture solution containing 2% newborn bovine serum and 3mmol of D-glucosamine salt, Incubate at 37°C. Harvest 72 hours after exposure, freeze and thaw for three times, centrifuge at 3000 r / min for 20 min, take the supernatant, and quantitatively divide it as porcine circovirus type 2 cytotoxicity.

[0036] The preparation method of PCV2 inactivated antigen liquid is as follows:

[0037] Add β-propiolactone (purity 97%) according to 1 / 4000 (V / V) of the total amount of virus solution, stir with addition, shake fully, inactivate at 4°C for 24 hours, and shake once every 4 hours. The inactivated virus liquid was hydrolyzed at 37° C. for 2 hours, and stored at 2-8° C. The inactivation test was qualified as PCV2 inactivated antigen liquid...

Embodiment 1

[0038] Embodiment 1, the method that examines porcine circovirus type 2 inactivated vaccine efficacy

[0039] 1 Vaccine preparation Dilute the PCV2 inactivated antigen solution with PBS to 10 6.5 TCID 50 / ml, 10 6.0 TCID 50 / ml and 10 5.5 TCID 50 / ml emulsified with adjuvant to prepare vaccines with different antigen contents for later use.

[0040] 2. Immunization of mice 40 Balb / c female mice were randomly divided into 4 groups, 10 mice in each group, immunized with PCV2 inactivated vaccine, each group was intraperitoneally injected with 0.2ml, and the immunization was boosted once with the same dose and the same inoculation method after 14 days. A non-immunized control group was established, and the immunization information is shown in Table 1. The sera of mice were collected 21 days after booster immunization for neutralizing antibody titer determination.

[0041] Table 1. Vaccine Immunization Information

[0042]

[0043] 3 Serum neutralizing antibody detection...

experiment example 1

[0065] Experimental example 1. Screening of judgment criteria for serum neutralizing antibody detection

[0066] (1) In addition, collect the mouse serum on the 21st day after the vaccine booster immunization in Example 1 for PCV2 neutralizing antibody titer determination. During the determination, steps 3.1-3.8 are the same as those in Example 1, but the determination standard in step 3.9 is: sample The number of PCV2-positive cells in the neutralization wells decreased by 80% compared with the number of positive cells in the virus control wells, and the neutralization efficacy was judged.

[0067] The serum neutralizing antibody levels of mice were detected under the above criteria, and the results were as follows:

[0068] The antibody levels of 10 mice in group 1# were 1:1084, 1:1331, 1:1184, 1:1347, 1:1184, 1:928, 1:994, 1:1255, 1:891, 1:1331 ; Antibody levels of 10 mice in 2# group were: 1:762, 1:600, 1:872, 1:502, 1:891, 1:994, 1:762, 1:802, 1:868, 1 :994;;The antibod...

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Abstract

The invention discloses a method for testing the efficacy of a porcine circovirus type 2 inactivated vaccine, and belongs to the technical field of biology. According to the method disclosed by the invention, serum is collected after a mouse is immunized with a vaccine, and the aim of testing the efficacy of the PCV2 vaccine is fulfilled by detecting the level of a PCV2 specific neutralizing antibody in the serum. According to the method for testing the efficacy of the porcine circovirus type 2 inactivated vaccine, the problems that time and labor are wasted during test pig screening, and the difference between batches of test results is large in a traditional piglet toxicity counteracting method are solved, and the steps of test pig dissecting and killing, virus separation and the like are avoided. The test method disclosed by the invention can truly reflect the immune effect of the porcine circovirus type 2 inactivated vaccine, is simple to operate, low in cost, short in time consumption, strong in sensitivity and high in stability, and has a wide application prospect in testing the efficacy of the porcine circovirus type 2 vaccine.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a method for testing the efficacy of porcine circovirus type 2 inactivated vaccine. Background technique [0002] Porcine circovirus belongs to the genus Circovirus of the family Circoviridae and is the smallest animal virus discovered so far. Porcine circovirus type 2 (PCV2) can cause multi-system functional disorders in pigs, such as piglet weaning failure syndrome, porcine dermatitis and nephrotic syndrome, porcine respiratory disease syndrome, etc. The main clinical manifestations are: pig physique decline, Diarrhea, weight loss, cough, wheezing, difficulty breathing, anemia and jaundice. After pigs are infected with porcine circovirus, it can lead to severe immunosuppression in pigs, especially piglets, which is prone to secondary or complicated other infectious diseases, mainly PRRS, swine fever (CSF), and Mycoplasma suis pneumonia. , Haemophilus parasuis disease,...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/68G01N33/569
CPCG01N33/6854G01N33/56983G01N2333/01G01N2469/20
Inventor 林艳刘汉平郑琴勤岳丰雄夏嘉鑫陈莉群杨勇袁雪林项聪英
Owner 成都史纪生物制药有限公司
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