Specific detection antigen of bovine echinococcosis granulosa and application of specific detection antigen

A technology of echinococcosis and antigen, which is applied in the field of medical devices, can solve problems such as cross-reaction, diagnostic error, and expensive preparation, and achieve the effects of strong specificity, rapid diagnosis, and high sensitivity

Active Publication Date: 2020-08-25
SHENYANG AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

These methods usually use natural echinococcus fluid, but the disadvantage of using natural echinococcus fluid is that it is prone to cross-reaction with sera from other diseased animals, expensive to prepare and difficult to commercialize
Although there have been many studies on the diagnosis of echinococcosis in humans, there are few studies on the diagnosis of animals, and when animals are naturally infected with the disease, the antibody response is not obvious, which easily leads to diagnostic errors

Method used

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  • Specific detection antigen of bovine echinococcosis granulosa and application of specific detection antigen
  • Specific detection antigen of bovine echinococcosis granulosa and application of specific detection antigen
  • Specific detection antigen of bovine echinococcosis granulosa and application of specific detection antigen

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Experimental program
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Embodiment 1

[0045] Embodiment 1, the preparation of Eg-H1 recombinant antigen

[0046] 1. Construction of pET-28a-Eg-H1 recombinant expression vector

[0047] In this embodiment, the nucleotide sequence of the Eg-H1 gene used is shown in SEQ ID NO.2, and the amino acid sequence of the protein is shown in SEQ ID NO.1. Eg-H1 is a multi-epitope recombinant antigen, and its epitopes are B-cell epitopes specific for two stages of Hexaconcus or Protoscoleia. Antigen B subunit 1, antigen B subunit 2, antigen Bsubunit 4, EG95, antigen protein, Ag5, EPC1, according to the bioinformatics analysis of their B cell epitope and secondary structure, intercepted some peptides and spliced ​​them into recombinant proteins , which was named Eg-H1.

[0048] Construct the pET-28a-Eg-H1 expression vector, which was synthesized by Shanghai Biotechnology Co., Ltd., and the pET-28a-Eg-H1 expression vector was verified by double enzyme digestion and sequencing. The double enzyme digestion steps are as follows: ...

Embodiment 2

[0097] Example 2, Preparation of Hypothetical protein EGR_01530 recombinant antigen

[0098] 1. Construction of pET-28a-Hypothetical protein EGR_01530 expression vector

[0099] In this example, the nucleotide sequence of the Hypothetical protein EGR_01530 gene used is shown in SEQ ID NO.4, and the amino acid sequence of the protein is shown in SEQ ID NO.3. Hypothetical protein EGR_01530 antigen is a hypothetical protein containing a cysteine ​​domain.

[0100] The prokaryotic expression vector pET-28a-Hypothetical protein EGR_01530 expression plasmid was constructed. The expression vector was synthesized by Shanghai Biotechnology Co., Ltd. The double enzyme digestion steps are as follows:

[0101] (1) Centrifuge the centrifuge tube containing the lyophilized powder of the expression vector of the target gene at 3000 rpm / normal temperature for 1 min.

[0102] (2) with 50 μL sterile ddH 2 O Dissolve the lyophilized powder, then mix gently with a vortex instrument, and centri...

Embodiment 3

[0109] Example 3, Preparation of bovine echinococcosis time-resolved fluorescent immunochromatographic test strips

[0110] Such as Figure 7 As shown, the time-resolved fluorescent immunochromatography test strip provided in this embodiment comprises a PVC base plate and a sample pad 2 positioned on the PVC base plate 1, a binding pad 3, a nitrocellulose film 5 (NC membrane, a chromatographic membrane), an absorbent paper 4. One end of the nitrocellulose membrane is connected to one end of the binding pad, the other end of the nitrocellulose membrane is connected to one end of the absorbent paper, one end of the sample pad is connected to the other end of the binding pad; the binding pad is coated with time-resolved fluorescently labeled Eg - For H1 recombinant antigen, a detection line 6 is set on the side of the nitrocellulose membrane close to the binding pad, a quality control line 7 is set on the layer close to the absorbent pad, and the detection line 6 is coated with ...

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Abstract

The invention discloses a specific detection antigen of bovine echinococcosis granulosa and application of the specific detection antigen. Specific detection antigen protein of the bovine echinococcosis granulosa is (a1) or (a2): (a1) protein of an amino acid sequence shown as SEQ ID NO.1, and (a2) protein obtained by carrying out substitution and / or deletion and / or addition of one or more amino acid residues on an amino acid sequence of the protein defined by the (a1) and with the same function. Time-resolved fluorescent microspheres are used as tracers, immunochromatographic test strips areprepared, the clinical operation is simple, the diagnosis is rapid, the sensitivity is high, and the specificity is high.

Description

technical field [0001] The invention relates to the technical field of medical devices, in particular to a specific detection antigen for bovine echinococcosis and its application. Background technique [0002] Echinococcus granulosus is a serious global zoonotic disease. Humans or ruminants mainly develop the disease by mistakenly eating the eggs excreted by Echinococcus granulosus, which is called "worm cancer". At present, there is little investment in research on diagnostic techniques for echinococcosis granulosus, and the most commonly used diagnostic method is to dissect and check the parasites. With the development of molecular techniques and immune techniques, there are now molecular diagnostics, such as LAMP, immunological diagnosis, Such as ELISA kit method, colloidal gold, etc., but it is found from clinical application that the detection accuracy of these methods is not high, the convenience is not strong, and it is not convenient for grassroots workers to operat...

Claims

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Application Information

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IPC IPC(8): C07K14/435C12N15/12G01N33/68G01N33/558
CPCC07K14/43554G01N33/558G01N33/6893G01N2333/43543
Inventor 杨娜陈启军姜宁丁莹莹桑晓宇冯颖陈冉王新一
Owner SHENYANG AGRI UNIV
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