Preparation method of avian infectious brunchitis virus HA antigen

A technology for bronchitis and chicken infectivity, applied in the field of bioengineering, can solve the problems of reduced potency, increased workload, protein denaturation, etc., and achieve the effects of prolonged storage period, reduced workload, and convenient storage

Inactive Publication Date: 2009-06-17
HENAN AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, most of the current HI detection methods still use live virus liquid as the diagnostic antigen. The main problems of this method are: (1) The live virus antigen has a large hidden danger, and there is a risk of loosening the virus; (2) The live virus antigen must be frozen. , th

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0021] (1) Virus proliferation

[0022] Will M 41 strain or HN 99 After the virus strain was diluted with sterile normal saline at a volume ratio of 1:100, inoculate 10-day-old SPF chicken embryos through the allantoic cavity, 0.1 mL per embryo, and continue to incubate at 37°C. Discard the dead embryos before 24 hours, and take them out after 36 hours. All embryos were placed in a 4°C refrigerator overnight, and the allantoic fluid was collected and stored at 4°C for later use.

[0023] (2) Virus liquid concentration and phospholipase C treatment

[0024] The collected fresh allantoic fluid was refrigerated and centrifuged at 3000rpm for 50min at a high speed, and the supernatant was centrifuged at a high speed of 25000rpm for 55min. The amount of HA buffer added was 1 / 200 of the volume of the supernatant obtained after the first centrifugation. Then add phospholipase C to the suspension, the final content is 1IU / mL, mix well and place in a 37°C water bath for 2h, shake o...

Embodiment 2

[0030] (1) Virus proliferation

[0031] Repeat the method for virus propagation in Example 1.

[0032] (2) Virus liquid concentration and phospholipase C treatment

[0033] The collected fresh allantoic fluid was refrigerated and centrifuged at 5000rpm for 40min at a high speed, and the supernatant was centrifuged at a high speed of 20000rpm for 60min. The amount of buffer added is 1 / 180 of the volume of the supernatant obtained after the first centrifugation; then add phospholipase C to the suspension, the final content is 1IU / mL, mix well and put it in a 37°C water bath for 2.2h, every Shake once every 20 minutes, take out the virus solution after the effect is completed, and store it at 4°C for later use;

[0034] (3) Antigen inactivation

[0035] Add the inactivator β-propiolactone according to 0.05% of the volume of the virus liquid, and inactivate it at 4°C for 24 hours;

[0036] (4) Antigen stabilization

[0037] The stabilizer sucrose was added according to the vo...

Embodiment 3

[0039] (1) Virus proliferation

[0040] Repeat the method for virus propagation in Example 1.

[0041] (2) Virus liquid concentration and phospholipase C treatment

[0042] The collected fresh allantoic fluid was refrigerated and centrifuged at 3500rpm for 48min at a high speed, and the supernatant was centrifuged at a high speed of 22000rpm for 62min. The amount of buffer added is 1 / 150 of the volume of the supernatant obtained after the first centrifugation; then add phospholipase C to the suspension, the final content is 1IU / mL, mix well and put it in a 37°C water bath for 2.5h, every Shake once every 20 minutes, take out the virus solution after the effect is completed, and store it at 4°C for later use;

[0043] (3) Antigen inactivation

[0044] Add the inactivator β-propiolactone according to 0.05% of the volume of the virus liquid, and inactivate it at 4°C for 24 hours;

[0045] (4) Antigen stabilization

[0046] The stabilizer polyethylene glycol is added accordin...

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PUM

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Abstract

The invention discloses a method for preparing a chicken infectious bronchitis virus HA antigen. The chicken infectious bronchitis virus HA antigen is prepared by the steps of virus multiplication, virus solution concentration and phospholipase C treatment, and antigen inactivation and stabilization. The stability of the prepared HA antigen at a temperature of 4 DEG C is prolonged to more than 6 months from the original 24 hours; besides, the valence of antigen is stable so as to solve the difficult problems which are not solved for a long time that the antigen is temporarily prepared for the test of each time and needs to be detected before use, and reduce the workload of the HI test. The prepared IBV HA antigen can be successfully applied to detecting the valence of antibody of serum HI after the chicken IBV vaccine immunization, and solves the problem that the detection of the IBV vaccine immunization effect is realized through neutralization tests in China for a long time, thereby wasting time and energy and having the danger of poison diffusion.

Description

technical field [0001] The invention belongs to the field of bioengineering, and in particular relates to a preparation process of chicken infectious bronchitis virus HA antigen. Background technique [0002] Chicken infectious bronchitis (Infectious bronchitis, IB) is an acute, highly contagious infectious disease caused by chicken infectious bronchitis virus (Infectious bronchitis virus, IBV). Symptoms include coughing, sneezing, and tracheal rales. The egg production of laying hens decreased, and deformed eggs were produced, and the performance of young chickens was particularly serious. Certain strains cause kidney and intestinal disease. Chicken infectious bronchitis has caused serious economic losses to the world poultry industry. Due to the high variability of IBV, there are many serotypes, and the cross-protective reaction between various serotypes is low, and the vaccine immunization often loses the expected effect. [0003] Most IBVs have no agglutinogen on the...

Claims

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Application Information

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IPC IPC(8): A61K39/215A61P31/12
Inventor 崔保安李新生高文明王莉王学斌杨明凡陈少渠冯利霞
Owner HENAN AGRICULTURAL UNIVERSITY
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