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Brucella fluorescence polarization (FPA) detection kit

A detection kit, Brucella technology, applied in the direction of measuring devices, immunoassays, instruments, etc., can solve problems such as difficult to cure, animal reproduction and production performance hazards

Inactive Publication Date: 2017-01-25
CHINA INST OF VETERINARY DRUG CONTROL
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

This disease not only has serious harm to the reproduction and production performance of animals, but more importantly, after people are infected with Brucella, it is often difficult to cure, thus causing serious public health problems.

Method used

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  • Brucella fluorescence polarization (FPA) detection kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0037] ——The quality standard of the bacterial species (B. suis S2 strain, B. suis S2) used for antigen preparation.

[0038] 1. Colony shape The edges of the colony are neat, round, dewdrop-shaped, irradiated by oblique light, and slightly blue opalescent when observed against the light.

[0039] 2. The staining form is cocci, scattered individually, without forming spores and capsules. The size is between 0.6 and 2.5 μm. Gram's stain was negative, Korotkoff's stain was red.

[0040] 3. The pure inspection is carried out according to the appendix of the existing "Chinese Veterinary Pharmacopoeia" (Chinese Veterinary Pharmacopoeia Committee, the People's Republic of China Veterinary Pharmacopoeia, 2010 edition three, China Agricultural Press, 2011, hereinafter referred to as "Chinese Veterinary Pharmacopoeia"), should purely.

[0041] 4. Specificity test

[0042] (1) Serological specificity Antigens made from cultures should agglutinate with smooth-type Brucella-positive s...

Embodiment 2

[0053] ——Purification and labeling method of the antigen OPS used in the detection of this application

[0054] 1. Bacterial suspension preparation Inoculate the qualified secondary seeds of B. suisS2 strain in pancreas agar flat bottles, and after culturing at 37°C for 48 hours, add 20ml of normal saline containing 0.5% phenol to each bottle to soak the surface of the bacteria for more than 10 minutes , wash the culture and collect in sterile glass bottles.

[0055] 2. Bacterial liquid inactivation and inactivation test Heat the collected bacterial suspension to 80°C for 120 minutes, and store it at 4°C after cooling naturally. The inactivation test should be carried out on the inactivated bacteria solution. Take 0.1ml of the inactivated bacteria solution and spread it on pancreatic agar or TSA plate, culture it at 37°C for 7 days, and it should not grow sterilely.

[0056] 3. Purification of OPS Centrifuge the bacterial suspension that passed the inactivation test at 10,00...

Embodiment 3

[0059] ——Preparation and inspection standards of negative control serum and positive control serum in the kit of this application

[0060] 1. Preparation of Negative Control Serum

[0061] (1) Animals are about 2-year-old, sexually mature healthy cattle that are negative for Brucella antibodies by tiger bengal plate agglutination test, test tube agglutination test and complement fixation test.

[0062] (2) Serum preparation Blood was collected from the jugular vein, the serum was separated, sterilized by filtration with a 0.45 μm filter, the collected serum was added with 0.05% proclin300 for preservation, and aseptically packaged, and after passing the inspection, it was stored at -20°C for later use.

[0063] 2. Test of negative control serum

[0064] (1) Appearance Orange or light yellow clear liquid.

[0065] (2) Sterility test is carried out according to the appendix of the current "Chinese Veterinary Drug Code", and the growth should be sterile.

[0066] (3) Specifici...

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Abstract

The invention relates to a Brucella fluorescence polarization (FPA) detection kit. The present invention uses fluorescein isothiocyanate (FITC) to mark Brucella glabrata O-chain lipopolysaccharide small molecule fragment (OPS) as an antigen. Based on the experimental principle of FPA, the kit can detect brucellosis in various animals, and can detect IgG and IgM antibodies in serum at the same time, so the kit has more obvious advantages in sensitivity; The improvement and optimization of the reagent formula, especially the precise calibration of the titer of the positive serum, effectively improved the sensitivity and specificity of the kit, and greatly shortened the sensitivity, repeatability, shelf life, and ease of operation. The time required for high-throughput detection, the invention provides a novel and efficient detection method for the diagnosis of brucellosis in my country.

Description

technical field [0001] The invention relates to a diagnostic method for animal brucellosis-brucella fluorescence polarization (FPA) detection kit, which belongs to the technical field of biological product detection. Background technique [0002] Brucellosis, referred to as brucellosis, is a zoonotic infectious disease mainly infecting domestic animals caused by Brucella. This disease not only has serious harm to the reproduction and production performance of animals, but more importantly, people are often difficult to cure after being infected with Brucella, thus causing serious public health problems. At present, the main way to eliminate the disease worldwide is to combine culling and immunization, so it is very necessary to establish a rapid and accurate diagnosis method for the prevention and elimination of brucellosis. [0003] Commonly used serological methods for the detection of Brucella antibodies include tiger red plate agglutination test (RBT), test tube aggluti...

Claims

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Application Information

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IPC IPC(8): G01N33/569
CPCG01N33/56911G01N2469/20
Inventor 蒋卉丁家波孙翠丽冯宇程君生朱良全彭小薇张磊
Owner CHINA INST OF VETERINARY DRUG CONTROL
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