Production method of fluorescently-labeled test strip for rapidly testing avian influenza virus

A bird flu virus and fluorescent labeling technology, which is applied in the field of animal disease detection, can solve the problems of difficult recording and storage of detection results, low detection sensitivity, and long time-consuming, etc., and achieve the effect of simple operation, high detection sensitivity and low detection cost

Inactive Publication Date: 2010-05-05
CHINESE ACAD OF INSPECTION & QUARANTINE
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AI Technical Summary

Problems solved by technology

Ordinary hemagglutination inhibition test (HI), enzyme-linked immunosorbent assay (ELISA), agarose diffusion test (AGID), immunofluorescence technique (IFT), gene amplification method (PCR), etc., all require certain experimental equipment And more professional technical operators, and it takes a long time
At present, the colloidal gold detection method is widely used in the initial screening of samples, but the disadvantage is that the detection sensitivity is low, it is difficult to carry out quantitative detection, and the detection results are not easy to record and save

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment Construction

[0005] 1. Materials

[0006] 50nm fluorescent nanoparticles, goat anti-mouse IgG antibody, avian influenza monoclonal antibody

[0007] Nitrocellulose membrane, glass cellulose membrane, absorbent paper, support plate

[0008] 2. Process flow

[0009] (1) Fluorescent nanoparticles with a size of 50nm suitable for protein labeling were prepared by reverse microemulsion method.

[0010] (2) Labeling of fluorescent nanoparticles and antibodies: Take 200 μL (0.5 mg / mL) of affinity-purified avian influenza monoclonal antibody A, dialyze with 0.025M carbonate buffer solution (pH 9.5) at 4°C for 6 hours; then add 100 μL to suspend Fluorescent nanoparticles (8mg / mL) in carbonate buffer solution, overnight at 4°C; add NaBH 3 CN, the final concentration is 5mM, react for 2h; add an equal volume of blocking solution (10mM tris 7.8, containing 2% BSA, 4% sucrose), and finally wash the labeled antibody 3 times with 10mM tris 7.8, and then wash with 100μL 10mM Tris 7.8 (containing 0.9% ...

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Abstract

A production method of a fluorescently-labeled test strip for rapidly testing avian influenza virus (AIV) belongs to the technical field of diagnosis and test methods of animal epidemic diseases. the test strip comprises a sample pad, a conjugate pad, a nitrocellulose membrane, an absorbent pad and a support plate. The sample pad, the conjugate pad, the nitrocellulose membrane and the absorbent pad are successively adhered to the support plate in turn. The conjugate pad is in overlap joint with the nitrocellulose membrane, and the nitrocellulose membrane is in overlap joint with the absorbent pad. An AI monoclonal antibody A, a fluorescent label is coated on the conjugate pad. A test line formed by an AI monoclonal antibody B and a quality control line formed by goat anti-mouse IgG are respectively coated on the nitrocellulose membrane. The test strip has the advantages of strong specificity, high sensitivity, short test time (20min), low test cost, simple and convenient operation and no need of professionals for operation, etc.

Description

technical field [0001] The invention relates to the technical field of animal epidemic detection methods, in particular to a method for manufacturing a fluorescent-labeled test strip for rapid detection of avian influenza virus. Background technique [0002] Avian influenza (Avian Influenza, AI), commonly known as fowl plague, is a kind of poultry infectious disease syndrome caused by type A influenza virus. The virus was first identified in chicken flocks in Italy in 1878. Now the disease spreads almost all over the world, and it poses a serious threat to the development of the poultry industry in the world, and even to human health. Avian influenza, especially highly pathogenic avian influenza (HPAI) of H5 and H7 subtypes, has been classified as Class A severe infectious disease by the International Veterinary Office (OIE), which has caused huge disasters in history. Ordinary hemagglutination inhibition test (HI), enzyme-linked immunosorbent assay (ELISA), agarose diffus...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/533G01N33/569
Inventor 邹明强张帆李锦丰
Owner CHINESE ACAD OF INSPECTION & QUARANTINE
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