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BEFV (Bovine Ephemeral Fever Virus) indirect ELISA (Enzyme Linked Immunosorbent Assay) antibody detection kit and preparation method thereof

A bovine epidemic fever virus and antibody detection technology, applied in the direction of measuring devices, instruments, scientific instruments, etc., can solve the problems of no application and large limitations in grassroots veterinary units and cattle farms, and achieve great operability and operation steps The effect of simplicity and loose technical requirements

Active Publication Date: 2014-08-06
LANZHOU INST OF VETERINARY SCI CHINESE ACAD OF AGRI SCI
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  • Abstract
  • Description
  • Claims
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Problems solved by technology

Therefore, it must be completed in a professional laboratory and operated by professionals. The limitations are very large, and there is basically no possibility of application in grassroots veterinary units and cattle farms.

Method used

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  • BEFV (Bovine Ephemeral Fever Virus) indirect ELISA (Enzyme Linked Immunosorbent Assay) antibody detection kit and preparation method thereof

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Embodiment Construction

[0018] 1. Antigen preparation

[0019] Design a pair of primers according to the selected gene fragment, the upstream primer sequence is 5'- catatg aatcattatgggatcggatc-3', downstream primer sequence is 5'- ctcgag tgtccatcttactgcctttg-3'. Those in italics and black in the primer sequences are endonuclease sites, and those in the upstream primers are Nde I restriction site, in the downstream primer is xho I restriction site. Use a viral RNA extraction kit to extract the BEFV RNA genome from BEFV-infected blood as a template, and use the above-mentioned primer pair to amplify the target DNA fragment by RT-PCR. The amplified target fragment is 480bp in length, located at 1141bp-1620bp of the BEFV G gene, and contains the G1 antigen epitope. Perform agarose electrophoresis on the amplified target DNA, purify and recover the target DNA, connect with the cloning vector pMD 19-T Simple, and transform into Escherichia coli DH5a; pick the positive monoclonal strain and extract ...

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Abstract

The invention relates to an indirect ELISA (Enzyme Linked Immunosorbent Assay) kit for detecting a BEFV (Bovine Ephemeral Fever Virus) serum antibody, as well as a preparation method of BEFV recombinant antigen used in the kit and the recombinant antigen. The recombinant protein only reacts with the BEFV antibody, and does not generate cross reaction with antibodies for other viruses in the same family, so that the recombinant protein has extremely high specificity; and the recombinant protein also has good reaction sensitivity. In addition, the repeatability of the kit is good, virus neutralization tests have high coincidence rate, and the storage life is long. The technical requirements are low during the operation of the kit, and the kit can be widely promoted and used in production, and applied to the BEFV epidemiological investigation and immune level monitoring.

Description

technical field [0001] The present invention relates to a detection kit for detecting mammalian diseases, specifically, the present invention is a detection kit for bovine ephemeral fever virus (BEFV) and a method for the detection kit. Background technique [0002] Bovine ephemeral fever (BEF) is an acute, febrile infectious disease of cattle, dairy cattle, beef cattle and buffalo caused by BEFV. BEF is transmitted by vector insects, and the relatively clear vectors are mosquitoes and Culicoides. The prevalence of this disease has obvious seasonality, often occurs in late summer and early autumn, and disappears in the first frost; the way of spreading is not affected by mountains and rivers, and it spreads in leaps and bounds. It can spread quickly, has a wide range of popularity, and has a certain periodicity. BEF occurs in Africa, Asia and Australia, and also occurs repeatedly in many provinces of my country, resulting in reduced milk production of dairy cows, decreased...

Claims

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Application Information

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IPC IPC(8): G01N33/569G01N33/535
CPCG01N33/54393G01N33/56983G01N2333/185
Inventor 殷宏郑福英刘志杰陈泽王积栋李志杨吉飞高闪电
Owner LANZHOU INST OF VETERINARY SCI CHINESE ACAD OF AGRI SCI
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