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Gelatin particle-agglutination detection reagent kit suitable for 1type duck hepatitis virus antibody and its uses

A duck hepatitis virus, latex agglutination technology, applied in the field of poultry immunology, can solve the problems of allergens (antigen or antibody shedding, short shelf life of latex reagents, difficult to be suitable for product development, etc. The effect of short, lightweight transport

Inactive Publication Date: 2011-07-20
HUAZHONG AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

When using latex agglutination technology to establish pathogenic detection methods, the specific antibody is generally sensitized to the surface of latex particles. The combination of ordinary polystyrene latex and antibody is non-selective physical electrostatic adsorption, and the sensitizer (antigen or antibody ) is easy to fall off from the latex particles or denatured and inactivated, resulting in a short shelf life of the sensitized latex reagent, which is difficult to meet the needs of product development

Method used

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  • Gelatin particle-agglutination detection reagent kit suitable for 1type duck hepatitis virus antibody and its uses
  • Gelatin particle-agglutination detection reagent kit suitable for 1type duck hepatitis virus antibody and its uses
  • Gelatin particle-agglutination detection reagent kit suitable for 1type duck hepatitis virus antibody and its uses

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] Example 1 Proliferation and Purification of Attenuated Virus of Type I Duck Hepatitis

[0031] 1. Proliferation of Type I Duck Hepatitis Attenuated Virus

[0032] The freeze-dried powder of the attenuated type 1 duck hepatitis strain (AV21111, purchased from the China Veterinary Drug Supervision Institute of the Ministry of Agriculture in Beijing) was diluted with sterilized normal saline at a volume ratio of 1:1000, and at the same time, 5% penicillin, strept After being treated with mycin for 1 hour, it was inoculated into the allantoic cavity of chicken embryos aseptically (0.1 mL / embryo). Then wax-sealed and placed in a 37.8°C incubator for culture. Eggs are illuminated 2 times a day. Chicken embryos that died within 24 hours were removed. The allantoic fluid of chicken embryos that died after 24 hours was aseptically collected for use.

[0033] 2. Purification of Type I Duck Hepatitis Attenuated Virus

[0034] Add 1% formalin solution to the chicken embryo all...

Embodiment 2

[0039] Example 2 Establishment of Anti-latex Agglutination Detection of Attenuated Type I Duck Hepatitis Virus

[0040] ① Latex pretreatment: Take 6ml of 10% polystyrene latex and put it into EP tube A, add 21ml of phosphate buffer (pH7.2, 0.01M). Weigh 0.3 g of trypsin and dissolve in 3 mL of phosphate buffer (pH 7.2, 0.01 M). , and then added to EP tube A in a water bath (56°C) for 18 hours, shaken 5-6 times in the middle, washed 3 times with carbonate buffer (pH9.6, 0.1M), and washed with carbonate buffer (pH9. 6, 0.1M) and resuspended to 30ml to prepare 2% latex, which can be stored at 4°C for later use.

[0041] ② Dilute the above 2% latex with carbonate buffer solution (pH9.6, 0.1M) to 1%, and use phosphate buffer solution (pH7.2 , 0.01M, adding 0.05% Tween) and washed twice. Then wash once with Tween-free phosphate buffer (pH7.2, 0.01M) and resuspend to 60 mL to obtain 1% sensitized latex. The specific steps are as follows:

[0042] 1. Determination of optimal sens...

Embodiment 3

[0068] Example 3 Preparation of Type I Duck Hepatitis Virus Antibody Positive and Negative Control Serum

[0069] 1 Animal preparation: Introduce 20 1-day-old ducklings without maternal antibodies, and keep them in isolation for 3 weeks. In the second week, take 5ml / feather duck blood from the wing vein to prepare negative serum, and drink water with 0.05% doxycycline to prevent bacterial infection. , Observation for a week, can be vaccinated when the performance is normal.

[0070] 2 Inoculation groups: A: 8 rats were inoculated with liver homogenate of virulent Zheng strain of duck hepatitis type I (preserved in our laboratory courtesy of Wuhan Institute of Virology, Chinese Academy of Sciences), B: 12 ​​rats were inoculated with allantoic fluid of attenuated strain of duck hepatitis I.

[0071] 3 Attenuated allantoic fluid treatment: Take 80ml of the collected allantoic fluid and freeze-thaw it twice, centrifuge at 5000rpm for 20min, take the supernatant and add an equal vo...

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Abstract

The invention belongs to the bird immunology technology area, it concretely relates to a kit of acceptable for the one type duck hepatitis virus antibody emulsion agglutination and the application of testing one type duck hepatitis virus antibody emulsion agglutination. The kit of this invention contains box body and core reagent in the box body, positive check sample, negative check sample, sample cell, scale sample sucker and the carrier of observed result. The characteristic is that core reagent is latex antigen from sublimating e duck hepatitis virus AV2111 and inactivating one type duck hepatitis virus sensitization polystyrene latex, the positive check sample is one type duck hepatitis virus antiserum, the negative check sample is the green duck of a day old of no one type duck hepatitis mother source antibody. The carrier is the paper characteristic latex agglutination card of having hollowness. The invention also discloses the preparation method and application of this kit. The kit has the significance merit of high specificity, high sensitivity, handy operation, and quickly diagnose.

Description

technical field [0001] The invention belongs to the technical field of poultry immunology. Specifically, the invention relates to the development of a latex agglutination kit suitable for type 1 duck hepatitis virus antibody and its application in the detection of type 1 duck hepatitis virus antibody. Background technique [0002] Duck virus hepatitis (DVH) is a highly lethal and highly transmissible viral infectious disease of ducklings caused by duck hepatitis virus (DHV). Clinically, it is characterized by obvious neurological symptoms and hepatomegaly with spot-like hemorrhage on the surface. It mainly affects ducklings less than 6 weeks old, especially ducklings 2 days to 3 weeks old, and adult ducks are resistant. After the onset of ducklings of different ages, the mortality rate is different, some are as high as 95%, and some are lower than 15%. The ducks that have endured become stiff ducks, and their growth and development are hindered and their economic benefits ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N33/576G01N33/543
Inventor 胡薛英彭婉芬程国富谷长勤张万坡
Owner HUAZHONG AGRI UNIV
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