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Immunocolloidal gold test paper for rapidly detecting mycoplasma bovis and preparation method thereof

A technology of colloidal gold test paper and mycoplasma bovis, which is applied in the field of immunology detection, can solve the problems of inability to realize rapid on-site detection, long detection time, cumbersome operation, etc., achieve fast detection speed, low detection cost, and improve detection efficiency.

Active Publication Date: 2019-03-29
CHONGQING UNIV OF TECH +1
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  • Abstract
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  • Claims
  • Application Information

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Problems solved by technology

[0005] For above-mentioned deficiencies in the prior art, the object of the present invention is to provide a kind of immunocolloidal gold test paper and preparation method thereof of quick detection Mycoplasma bovis, solve the existing Mycoplasma bovis detection method to have complicated operation, long detection time, can not realize on-the-spot rapid detection and high cost

Method used

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  • Immunocolloidal gold test paper for rapidly detecting mycoplasma bovis and preparation method thereof
  • Immunocolloidal gold test paper for rapidly detecting mycoplasma bovis and preparation method thereof
  • Immunocolloidal gold test paper for rapidly detecting mycoplasma bovis and preparation method thereof

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Embodiment

[0047] 1. Antigen screening

[0048] The cell membrane surface protein of Mycoplasma bovis plays an important role in the process of pathogenicity and infection. The P33 gene was screened out by predicting the signal peptide and epitope analysis of the cell membrane surface protein of Mycoplasma bovis, and the bioinformatics of the P33 gene and its encoded protein were further carried out. The results of biological and immunological analysis prove that the P33 protein is a specific adhesion protein on the cell membrane of Mycoplasma bovis.

[0049] 2. Prokaryotic expression and detection of Mycoplasma bovis P33 protein

[0050] 2.1 Construction of pET28a(+)-P33 recombinant vector

[0051] According to the gene sequence of P33 (accession number AIA33909.1) published in GenBank, without changing the amino acid sequence of P33 protein, the gene sequence was optimized according to the codon preference of Escherichia coli, and a pair was designed using Primer Premier5.0 software. ...

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Abstract

The invention discloses an immunocolloidal gold test paper for rapidly detecting mycoplasma bovis and a preparation method thereof. The bottom layer of the test paper is a support plate, the support plate is sequentially adhered with a sample pad, a bonding pad, an NC film and an absorption pad which are closely connected. A mycoplasma bovis P33 monoclonal antibody with a colloidal gold mark is absorbed on the bonding pad, a rabbit anti-mycoplasma bovis P33 polyclonal antibody is sprayed on the NC film to take as a detection line and a sheep anti-BALB / C mouse IgG antibody is sprayed on the NCfilm to take as a quality control line, the line widths of the detection line and the quality control line are 1mm, the distance of the detection line and the quality control line is 5mm, the detection line and the quality control line are located at the center of the NC film, and the amino acid sequence of the mycoplasma bovis P33 protein is as shown such as SEQ ID NO.1. The test paper can be used for rapid detection of the mycoplasma bovis antigen to generate a result after 5-10min, the sensitivity is 5 ng / mL, the sample amount is few, cross reaction is not needed, the flexibility is high, the requirements of low cost and rapid field diagnosis can be met, and the detection efficiency can be greatly improved. The immunocolloidal gold test paper for rapidly detecting mycoplasma bovis and the preparation method thereof can develop important effects in the prevention and control of bovine mycoplasmosis and have potential promotion and application values.

Description

technical field [0001] The invention relates to the technical field of immunological detection, in particular to an immune colloidal gold test paper for rapidly detecting mycoplasma bovis and a preparation method thereof. Background technique [0002] Mycoplasma bovis (Mycoplasma bovis, Mb) is a kind of simple and extremely small prokaryotic microorganisms without cell walls, between bacteria and viruses, capable of self-replication and independent living, which can cause bovine pneumonia, mastitis, arthritis and many other diseases. This disease has caused huge economic losses and seriously threatened the healthy development of my country's cattle industry. [0003] At present, the detection of Mycoplasma bovis in the prior art mainly includes methods such as pathogen isolation and identification, polymerase chain reaction (PCR), fluorescent quantitative PCR technology, enzyme-linked immunosorbent assay (ELISA), but these methods are all subject to clinical application. Lar...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/532G01N33/558G01N33/68
CPCG01N33/532G01N33/558G01N33/68
Inventor 吴胜昔梁望旺曾政李令臣李俊萱侯力嘉鲁友铭蔺露陈忠琼
Owner CHONGQING UNIV OF TECH
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