Novel immunochromatography test paper for detecting human brucellosis antibody and preparing method thereof

A Brucella and human detection technology, which is applied in the field of immunological diagnosis and detection, can solve the problems of lack of detection equipment and technical personnel, and achieve the effects of high sensitivity, long shelf life and strong sensitivity

Inactive Publication Date: 2015-02-18
SHIHEZI UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] In my country's remote areas, grassroots medical institutions and animal husbandry and veterinary stations lack professional testing equipment and technical personnel. Therefore, it is particularly important to develop a simple, economical, convenient and fast detection method for the control and treatment of the disease.

Method used

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  • Novel immunochromatography test paper for detecting human brucellosis antibody and preparing method thereof
  • Novel immunochromatography test paper for detecting human brucellosis antibody and preparing method thereof
  • Novel immunochromatography test paper for detecting human brucellosis antibody and preparing method thereof

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] Embodiment 1: Prokaryotic expression and purification of Brucella outer membrane protein BP26

[0027] 1.1 Design and synthesis of primers

[0028] Referring to the VirB5 gene sequence (accession number: BMEII0029) of the Brucella melis reference strain 16M published on GenBank, according to the characteristics of the pGM-T cloning vector and the pET-28a prokaryotic expression vector, both ends were designed to contain restriction internal Primers for Dicer Xho I / EcoR I restriction sites. See Table 1-1 for primer sequences.

[0029] Table 1-1 Primer sequences

[0030] Table 1-1 Primer sequences

[0031]

[0032] 1.2 PCR amplification of the target fragment

[0033] PCR amplification was performed using the Brucella 16M genome as a template, and the PCR reaction system is shown in Table 1-2. The reaction conditions used were: 94°C pre-denaturation for 5 min; 95°C for 50 s, 58°C for 45 s, 72°C for 1 min, 30 cycles; 72°C extension for 7 min; 4°C. After the reactio...

Embodiment 3

[0064] Embodiment 3: the colloidal gold immunochromatography test strip of Brucella antibody in the preparation serum

[0065] 1.1 Preparation of colloidal gold

[0066] Clean the triangular flask for burning colloidal gold and then soak it in the acid tank overnight; take it out and rinse it with clean water for 20 times, then wash it with distilled water for 3 to 5 times, and then rinse it with ultrapure water for 3 times. After drying, it is treated with a silylating agent, rinsed with ultrapure water after drying, and the triangular flask is dried to prepare colloidal gold.

[0067] Add 50ml of chloroauric acid with a concentration of 0.01% into the above-mentioned treated conical flask, heat and stir on a magnetic stirrer, and quickly add 1ml of trisodium citrate aqueous solution with a concentration of 1% with a pipette after the solution boils, the color of the solution It turns into a stable wine red in a short period of time. After cooling, it is restored to 50ml wit...

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Abstract

The invention relates to the field of zoonosis immunologic diagnosis and discloses a brucellosis antibody test strip and a preparing method thereof. The brucellosis antibody test strip is prepared in a way that colloidal gold with a grain diameter of 30nm is first prepared, used as an indication mark for marking and reconstructing large peptostreptococcus proteins L, dissolved in a gold mark work solution and sprayed on glass fibers to make a gold mark pad; VirB5 genes are cloned from a brucella genome and connected to pET-28a to form an expression vector; the expression vector is converted into escherichia coli, VirB5 proteins are expressed and coat on a nitrocellulose membrane after separation and purification as a detection line, IgGs (Immunoglobulin G) bonded with the proteins L are coated on the nitrocellulose membrane as a quality control line, and an immunochromatography detection device is made. The brucella immunochromatography device has the characteristics of strong specificity, high sensitivity, great stability, simplicity, economy, rapidness and the like and has extremely significant actual application value in the monitoring and epidemic control of zoonosis.

Description

technical field [0001] The invention belongs to the technical field of immunological diagnosis and detection, and in particular relates to a test paper for detecting antibodies produced in vivo after human infection with Brucella and a preparation method thereof. The invention also relates to the genetic engineering preparation process of VirB5, which is one of Brucella type IV protein secretion system family, as a diagnostic antigen, and the application of the protein in the detection of Brucella antibody. technical background [0002] Brucellosis (brucellosis, referred to as brucellosis) is a "reemerging" zoonotic infectious disease that seriously endangers the health of humans and livestock. class of infectious diseases. The disease is prevalent in about 170 countries and regions around the world. In recent years, the human and animal epidemics of brucellosis have shown a trend of recovery around the world, showing a continuous growth trend and intensifying. In my coun...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/569
CPCG01N33/56911G01N33/6893G01N2333/23
Inventor 石峰王远志陈创夫程婷婷钱红李志强
Owner SHIHEZI UNIVERSITY
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