Fast-detection test strip of bluetongue virus (BTV) and preparation method

A bluetongue virus and detection test paper technology, applied in the biological field, can solve the problems of poor specificity of AGID, and achieve the effect of rapid detection, high sensitivity and strong specificity

Inactive Publication Date: 2009-12-09
花群义
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  • Abstract
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  • Claims
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Problems solved by technology

Among BLU detection methods, Agar Gel Immuno Diffusion (AGID) has been widely used in the world because of its simple operation and low cost. It is also one of the methods recommended by OIE so far. However, the antigen used in this method There is a certain cross-reactivity with related viruses such as Epizootic hemorrhagic disease virus of deer (EHDV), so the specificity of AGID detection is poor

Method used

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  • Fast-detection test strip of bluetongue virus (BTV) and preparation method

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Embodiment Construction

[0028] 1. Preparation process of monoclonal antibody against bluetongue virus VP7 protein:

[0029] (1) After BHK-21 cells grow into a monolayer, BTV is inoculated and reacted at 37°C for 1 hour. Serum-free basal medium was added to continue culturing, and the virus was harvested when the cytopathic pathology (CPE) reached more than 75% after 2-3 days. The harvested virus liquid was repeatedly frozen and thawed 3 times, centrifuged at 8000r / min for 30min, and the supernatant was taken. Ultracentrifuge at 28000r / min at 4°C for 3h. The precipitate was dissolved with 1 / 100 of the original volume of PBS, and then 20%, 60% (w / v) discontinuous sucrose density gradient 20,000r / min was used for ultracentrifugation at 4°C for 3h, and the purified virus band was harvested as an immune antigen, and DU800 nucleic acid was used to The protein content was measured by a protein analyzer and stored at -20°C for later use.

[0030] (2) After the first immunization of BALB / c mice with the pu...

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Abstract

The invention relates to the biotechnology field. A fast-detection test trip of a bluetongue virus (BTV) comprises a sample absorption pad 1, a nitrocellulose membrane 3, an absorbent pad 6 and a baseplate 7; wherein, the sample absorption pad, the nitrocellulose membrane and the absorbent pad are fixed on the baseplate by arrangement in sequence; the sample absorption pad is a porous fiber which is linearly arranged and fixed on an impermeable material; the sample absorption pad is loaded with a colloidal gold pad which is a conjugate of a monoclonal antibody of a BTV VP7 protein and the colloidal gold; a detection line 4 is positioned on the nitrocellulose membrane and a polyclonal antibody of the BTV is coated or sprayed on the detection line; a control line 5 is positioned on the nitrocellulose membrane and a protein A is coated or sprayed on the control line. The test strip of the invention features strong specificity, high sensitivity and fast detection, can show results in 10 minutes and is especially suitable for port quarantine, field and on-site quarantine. The test strip can detect the BTV antigen directly and rapidly, without special equipments or technicists.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a rapid detection test strip for detecting bluetongue virus antigen and a preparation method of the rapid detection test strip. Background technique [0002] Bluetongue disease (BLU) is a severe animal infectious disease caused by bluetongue virus (BTV) of the reoviridae family (reoviridae) orbivirus genus (Obivirusgenus), and is transmitted by Culicoides , mainly harm sheep, goats, cattle and other livestock, and is widely prevalent in the world. The World Organization for Animal Health (OIE) lists BLU as an important animal infectious disease. In many national animal trade agreements, BLU is one of the infectious diseases that must be quarantined. In recent years, with climate warming, the incidence of BLU has increased in many European countries, seriously affecting international animal trade. [0003] There are 24 serotypes of BTV, and the BTV genome contains 10 double-stranded...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/569G01N33/577G01N33/558G01N33/532G01N33/552
Inventor 花群义杨俊兴阮周曦曾少灵曹琛福吕建强陈兵秦智锋卢体康杨建明林庆燕陶虹
Owner 花群义
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