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Porcine reproductive and respiratory syndrome virus antibody detection kit

A respiratory syndrome and antibody detection technology, applied in biological testing, measuring devices, material testing products, etc., can solve problems such as poor repeatability, poor sensitivity and specificity of PRRS ELISA kits, and limited wide application

Inactive Publication Date: 2016-05-25
SHANDONG BINZHOU ANIMAL SCI & VETERINARY MEDICINE ACADEMY
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  • Abstract
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Problems solved by technology

However, due to its high price (the kit priced at ￥5500 for 2×96-well plates and ￥13000 for 5×96-well plate kits), its wide application in the domestic market was greatly limited.
[0003] In addition, most of the PRRSELISA kits independently developed and produced in China have problems such as poor sensitivity, specificity, and poor repeatability.

Method used

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  • Porcine reproductive and respiratory syndrome virus antibody detection kit
  • Porcine reproductive and respiratory syndrome virus antibody detection kit
  • Porcine reproductive and respiratory syndrome virus antibody detection kit

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Effect test

Embodiment 1

[0058] The present invention establishes pET-777 protein as the ELISA method of coated antigen through the following steps:

[0059] 1. Isolation and identification of PRRSVDY strain

[0060] 1. Virus isolation and virus passage

[0061] The diseased pig tissue and lung samples freshly collected from a pig farm in Dongying were aseptically cut into pieces, ground with normal saline, the suspension was sucked, frozen and thawed three times, centrifuged at 12,000rpm for 10 minutes, and the supernatant was stored at -20°C.

[0062] Dilute the treated sample to a 1:10 suspension with DMEM maintenance medium containing penicillin and streptomycin, filter and sterilize at 0.22 μm, take 1 ml to inoculate a single layer of Marc-145 cells, absorb at 37°C for 1 hour, add maintenance The virus solution was continuously cultivated, and the cell lesion was observed. When 75% of the cells were lesioned, the virus liquid was collected and continued to be passed on.

[0063] Through subcult...

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Abstract

The invention discloses a porcine reproductive and respiratory syndrome virus antibody detection kit and belongs to the technical field of biological detection. The detection kit comprises a porcine reproductive and respiratory syndrome virus ELISA antigen coating plate, and the antigen in the ELISA antigen coating plate is protein expressed and purified by a recombinant expression plasmid pET777 obtained through double digestion of the gene segment NSP7-777 of a PRRSV DY strain and pET-32a by means of restriction enzymes BamHI and SacI. An antibody can be detected within three days after PRRSV DY strain NSP7 protein is immunized, and a high antibody level can still be realized within 248 days. The coincident rate of the detection method and an IDEXX kit can reach 99% or above, however, the cost is only one eighth that of the IDEXX kit. The porcine reproductive and respiratory syndrome virus antibody detection kit can replace imported kits to be used and popularized.

Description

technical field [0001] The invention relates to the technical field of biological detection, in particular to a porcine reproductive and respiratory syndrome virus antibody detection kit. Background technique [0002] At present, the quality of domestic PRRSV antibody detection kits is uneven. Among them, the American IDEXXPRRSELISA kit is widely recognized worldwide. However, due to its high price (the kit with 2×96-well plates is priced at ￥5500, and the kit with 5×96-well plates is priced at ￥13000), its wide application in the domestic market is greatly limited. [0003] In addition, most of the PRRSELISA kits independently developed and produced in China have problems such as poor sensitivity, specificity, and poor repeatability. [0004] Enzyme-linked immunosorbent assay (ELISA) is suitable for grass-roots veterinary work because of its low condition requirements, simple operation, good stability, and high specificity and sensitivity. It has been widely used in the d...

Claims

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Application Information

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IPC IPC(8): G01N33/569G01N33/68
CPCG01N33/56983G01N33/68
Inventor 张松林刘鸿艳李峰任艳玲刘磊马永彪沈志强
Owner SHANDONG BINZHOU ANIMAL SCI & VETERINARY MEDICINE ACADEMY
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