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Avian influenza virus latex agglutination assay kit and its use

An avian influenza virus and latex agglutination technology, applied in the field of avian immunology, can solve the problems of difficulty in product development, short shelf life of latex reagents, and difficulty in binding to latex surfaces, etc., and achieves a wide range of detection samples, short detection time, and easy operation. Effect

Active Publication Date: 2010-11-17
WUHAN KEQIAN BIOLOGY CO LTD
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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, it is generally to sensitize specific antibodies to the surface of latex particles. The combination of ordinary polystyrene latex and antibodies is non-selective physical electrostatic adsorption. Due to the small molecular weight of antibodies, it is very difficult to Difficult to bind to the latex surface, even the sensitized 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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  • Avian influenza virus latex agglutination assay kit and its use
  • Avian influenza virus latex agglutination assay kit and its use
  • Avian influenza virus latex agglutination assay kit and its use

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0037] The specific separation and identification steps are as follows:

[0038] (1) Incidence and necropsy lesions of sick ducks submitted for inspection

[0039] The egg production of laying ducks decreased rapidly (dropped to 10%), and small eggs and deformed eggs were produced. Necropsy showed follicular atrophy, some hyperemia and hemorrhage. Breeding ducklings began to get sick at the age of 20 days, with decadent diet, or even waste, and white loose feces. The respiratory symptoms were not typical, and the main manifestations were neurological symptoms. Previous reports on the symptoms of the same subtype of avian influenza strains were quite different. After 6 days, the mortality rate reached its peak, and the daily death rate was above 5%. The autopsy found that there were hemorrhage points in the coronary groove fat of the heart, and a large area of ​​gray-white strip necrosis appeared in the myocardium, which is also a typical lesion of the diseased duck. Althoug...

Embodiment 2

[0089] (1) Proliferation of virus

[0090] Get QLG-1 virus strain, make 10 with sterilized normal saline -4 Dilute and inoculate specific pathogenic microorganism-free (SPF) chicken embryos into the allantoic cavity, 0.2 mL per embryo, seal the pinhole after inoculation, and continue to incubate at 37°C. Eggs were illuminated once every 12 hours, and the chicken embryos that died 24 hours ago were discarded. The chicken embryos that died 24 hours to 72 hours were cooled at 4°C overnight, and the allantoic fluid of chicken embryos was harvested. The HA titer ≥ 9 log2, the formula is qualified.

[0091] (2) Inactivation of virus

[0092] After freezing and thawing the qualified chicken embryo allantoic fluid for 3 times, add formaldehyde with a final concentration of 0.2%, mix well and replace it into another sterile container, act at 37°C for 24 hours, shake every 6 hours for 1 Second-rate. The allantoic fluid of inactivated chicken embryos was sampled to inoculate 9-11-day...

Embodiment 3

[0105] 1 Determination of the amount of optimally coupled IgG

[0106] Add different amounts of IgG to 1 mL of carboxylated latex suspended in boric acid buffer, increasing from 200 μg, 400 μg, and 600 μg to 1800 μg. Protein amount, calculate the coupling efficiency of IgG, the formula is as follows:

[0107] Coupling efficiency of IgG (%)=(Initial amount of IgG added-remaining amount of IgG) / Initial amount of IgG added×100%

[0108] It has been verified by experiments that in 1 mL of carboxylated latex with a concentration of 1%, when the amount of IgG added is 800-1000 μg, the coupling efficiency reaches the maximum, and at this time the latex detection has the highest sensitivity and the best stability. The specific results are shown in Table 8:

[0109] The coupling efficiency when table 8 is different IgG adding amount

[0110]

[0111] 2 Selection of the best coupling time

[0112] Add 1000 μg IgG to 1 mL of carboxylated latex suspended in boric acid buffer, and r...

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Abstract

The invention discloses a reagent box of latex agglutination which is used for quickly detecting the birds flu virus. Putting the hare birds flu anti-virus nuclein IgG couple to the surface of carboxylation polystyrene latex particles, founding the method to test the latex agglutination of birds flu virus, then supplemented by other match reagents prepared the reagent box of detect the bird flu virus latex agglutination. the reagent box of detect the bird flu virus latex agglutination is composed of box body and latex diagnostic reagents, sample handling liquid of A which is located in box, sample handling liquid of B, positive contrast sample and negative contrast sample. The reagent box is referred directly detect the birds flu virus, its notable advantages are that strong specificity, high sensitivity, simple operation and rapid diagnosis.

Description

technical field [0001] The invention belongs to the technical field of poultry immunology, and relates to related fields such as immunology and applied chemistry. Specifically, the present invention relates to a latex agglutination detection kit of avian influenza virus and the application of the kit in detection of avian influenza virus. Background technique [0002] As we all know, avian influenza is a severe infectious disease that seriously endangers the poultry industry and endangers human health. The "Hong Kong bird flu" incident in 1997 was the first direct cross-species transmission of bird flu to humans, and the Southeast Asian bird flu in 2004 aroused great concern around the world on the cross-species spread of bird flu. In my country in 2004, there were also 50 epidemic spots in 16 provinces and cities where bird flu outbreaks broke out on a large scale, which caused great losses to the poultry industry in my country. [0003] At present, the pathogenic methods...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N33/569G01N33/544
Inventor 金梅林陈剑锋喻正军张安定李红超陈焕春
Owner WUHAN KEQIAN BIOLOGY CO LTD
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