Animal brucella antibody colloidal gold test paper film detection reagent kit

A technology of colloidal gold test paper and Brucella, which is applied in the field of animal epidemic diagnosis, can solve the problems of time-consuming, laborious, and inability to distinguish the pathogenic types of Brucellosis, and achieve the effect of rapid detection

Inactive Publication Date: 2013-01-09
MILITARY VETERINARY RES INST PLA MILITARY MEDICAL ACAD OF SCI
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  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

They all have the following deficiencies. First, it is difficult to avoid cross-reaction with other bacteria (such as: Yersinia O9, Escherichia coli O157); second, traditional diagnostic methods are time-consuming, laborious and prone to false positives; Third, traditional diagnostic methods cannot distinguish the pathogenic types of brucellosis

Method used

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  • Animal brucella antibody colloidal gold test paper film detection reagent kit
  • Animal brucella antibody colloidal gold test paper film detection reagent kit

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Experimental program
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Effect test

Embodiment 1

[0032] 1. Extraction and identification of animal Brucella LPS

[0033] The LPS of Brucella bovis 544A and Brucella sheep 16M were extracted by hot phenol method and cold phenol method respectively, and purified by Sephadex G-50 chromatography column, and the purity and immunological characteristics of the purified LPS were tested. Identification. The results showed that the content of LPS was 1.21mg / mL-2.31mg / mL, and the purity was 96.1%-97.2%. It only reacted with the corresponding antibody, and there was no cross-reaction between LPS of different strains.

[0034] 2. Preparation of colloidal gold and selection of optimum particle size

[0035] Use sodium citrate reduction method and tannic acid-sodium citrate reduction method to prepare 20nm, 25nm, 30nm, 40nm colloidal gold particles, observe the dispersion and uniformity results under the electron microscope, and then select the most suitable colloidal gold particle size of 20nm and 25nm to mark.

Embodiment 2

[0037] 1. Determination of the optimal labeling pH of the animal Brucella gold standard probe

[0038] Using colloidal gold gradient method and O value curve method, the optimum pH value of colloidal gold-labeled SPA was determined to be 5.2-6.8.

[0039] 2. Determination of the optimal labeling amount of the animal Brucella gold standard probe

[0040] Using the colloidal gold gradient method and the O value curve method, the optimal labeling amount of colloidal gold-labeled SPA was determined to be 20-60 μg / mL.

[0041] 3. Preparation and purification of gold-labeled probes

[0042] Take two test tubes, add 5mL 20nm and 25nm colloidal gold respectively; add 30μ1~50μl0.2mol / L K 2 CO 3 , adjust the pH value to 5.2~6.8; calculate the total amount of protein to be labeled according to the total amount of colloidal gold used for labeling to be 60μg / ml~90μg / ml, and add SPA to the colloidal gold solution drop by drop , the whole process is about 5min. Shake well for 15 minutes...

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Abstract

The invention discloses an animal brucella antibody colloidal gold test paper film detection kit, which bases on immunology basic principle of antigen and antibody specific combination, utilizes colloidal gold immunity chromatography technique and silver dye reinforcement technique, uses combined golden yellow staphylococcus A protein (SPA) as gold marking antibody, uses high purity brucella LPS as peridium antigen, and utilizes goat anti-bovine IgG as quality control line to prepare colloidal gold test paper tape. The invention can accurately differentiate species of animal brucella and can synchronously detect animal brucella disease of cattle, sheep and pig species with no crossing reaction and low false negative proportion, negative and positive coincidence ratio of the invention and conventional separation and culture is 100%, detection result can be obtained in 5 min, and the kit need no assistant complicated detection instrument.

Description

technical field [0001] The invention discloses an animal Brucella antibody colloidal gold test paper film detection kit, which is used for the diagnosis of brucellosis and belongs to the technical field of animal epidemic diagnosis. Background technique [0002] Brucella is a Gram-negative, specific intracellular parasite that can infect a variety of animals and humans. At present, Brucella is divided into 6 species according to the pathogenicity and host, namely, Brucella sheep, Brucella bovis, Brucella pig, Brucella sylvania, and Brucella ovis. bacteria, Brucella canis. Among them, brucella of cattle, sheep, and pigs are widely prevalent in the world, which is extremely harmful to animal husbandry. More importantly, they can cause human infection and threaten public health. Therefore, the research on the diagnosis of brucellosis has always been concerned by countries all over the world. Traditional serological diagnostic methods for diagnosing animal brucellosis include...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N33/569G01N33/558
Inventor 王兴龙李晓艳唐景峰梅建军
Owner MILITARY VETERINARY RES INST PLA MILITARY MEDICAL ACAD OF SCI
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