Echinococcosis diagnosis immunochromatography test strip based on circulating antigen detection

An echinococcosis and antigen technology, which is applied in the field of immunochromatographic test strips for the diagnosis of echinococcosis, can solve the problems of not being able to reflect the infection status of echinococcosis, and achieve clear and intuitive results, high sensitivity and specificity, and stability good sex effect

Pending Publication Date: 2022-02-01
中国疾病预防控制中心寄生虫病预防控制所国家热带病研究中心
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Problems solved by technology

[0005] The purpose of the present invention is to propose an immunochromatographic test strip and preparation method for diagnosing echinococcosis based on the detection of circulating antigens, to solve the problem that the existing immunodiagnostic technology cannot reflect the infection status of echinococcosis, and for the early stage of echinococcosis Diagnosis, curative effect evaluation, prognosis judgment and epidemiological investigation are also of great significance

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  • Echinococcosis diagnosis immunochromatography test strip based on circulating antigen detection
  • Echinococcosis diagnosis immunochromatography test strip based on circulating antigen detection
  • Echinococcosis diagnosis immunochromatography test strip based on circulating antigen detection

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Embodiment 1

[0032] The preparation of the monoclonal antibody of embodiment 1 echinococcosis cyst fluid antigen

[0033] Preparation of hydatid disease purified cyst fluid antigen: take fresh or frozen sheep liver cyst fluid and centrifuge at 3000rpm for 30 minutes to remove the precipitate. The supernatant was dialyzed overnight against 5 mM acetate buffer solution with pH 5.0, and the dialysate was centrifuged at 5000 rpm for 30 minutes to collect the precipitate. Dissolve the precipitate with 200mM PBS at pH 8.0, then add saturated ammonium sulfate to 40%, let stand at 4°C for 12 hours; centrifuge at 13000rpm for 30 minutes to remove the precipitate, and dialyze the supernatant against pH 7.4, 10mM PBS overnight. The dialysate was recovered and the protein concentration was measured to obtain the purified cystic fluid antigen of echinococcosis. This antigen is used to prepare monoclonal antibodies.

[0034] Immunization: Each BALB / c mouse was injected intraperitoneally with 100 μg of...

Embodiment 2

[0045] Example 2 Preparation of Immunochromatographic Test Strips for Diagnosis of Echinococcosis Based on Detecting Circulating Antigens

[0046] 1. The monoclonal antibody against hydatid cyst fluid antigen was prepared from Example 1.

[0047] 2. Goat anti-mouse IgG was purchased from Cohesion Biosciences.

[0048] 3. Preparation of Monoclonal Antibody Colloidal Gold Probe Solution

[0049] (1) Colloidal gold is prepared by citric acid reduction method:

[0050] The HAuCl4 (Shanghai Sinopharm Group Chemical Reagent Co., Ltd.) aqueous solution that is 0.01% by mass percent concentration is boiled, and 2 mL of trisodium citrate aqueous solution that is 1% by mass percent concentration is added under stirring, and continues to boil until the liquid is dark red to obtain colloidal gold solution.

[0051] (2) Preparation of immune colloidal gold probe pad:

[0052] with 0.1M K 2 CO 3 Adjust the pH value of the solution to 7.2, add anti-hydatid cyst fluid antigen monoclonal...

Embodiment 3

[0061] Example 3 Laboratory and on-site detection and evaluation of immunochromatographic test strips for the diagnosis of echinococcosis based on detection of circulating antigens

[0062] 1. Detection method

[0063] Sample requirements: The test sample is whole blood or serum, using a blood collection needle to collect peripheral blood (on-site) or using collected venous anticoagulant blood (laboratory). The sample should be tested as soon as possible after collection. If it cannot be tested in time, the collected whole blood sample needs to wait for its natural agglutination to collect serum, which can be stored at 2-8°C for 3 days. If long-term storage is required, it should be stored below -20°C. Avoid repeated freeze-thaw cycles. Samples need to be shipped in vacuum flasks or other devices containing dry ice.

[0064] Detection method: Use a capillary blood collection tube to draw the blood sample to be tested to the scale line (5μL), and put it into the test strip de...

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Abstract

The present invention provides an echinococcosis diagnosis immunochromatography test strip based on circulating antigen detection. the echinococcosis diagnosis immunochromatography test strip comprises a sample pad, a colloidal gold probe pad, a cellulose membrane and a water absorption pad, the cellulose membrane is provided with a detection line and a quality control line, the colloidal gold probe pad comprises a monoclonal antibody which is labeled by colloidal gold and is specifically combined with an echinococcosis cyst fluid antigen, and the monoclonal antibody on the colloidal gold probe is generated by a mouse hybridoma cell with the preservation number of CGMCC (China General Microbiological Culture Collection Center) No.22389; and one detection line comprises a monoclonal antibody specifically combined with an echinococcosis cyst fluid antigen, and the monoclonal antibody on the detection line is generated by mouse hybridoma cells with the preservation number of CGMCC (China General Microbiological Culture Collection Center) No.22388. The echinococcosis diagnosis immunochromatography test strip has the advantages of simplicity, sensitivity, specificity and rapidness, and is suitable for clinical and on-site use.

Description

technical field [0001] The invention belongs to the field of bioengineering, and relates to a colloidal gold immunochromatographic test strip, in particular to an immunochromatographic test strip for diagnosing echinococcosis based on detecting circulating antigens. Background technique [0002] Echinococcosis, also known as Hydatid disease or Hydatidosis, is a zoonotic parasitic disease caused by the larval stage of tapeworms of the genus Echinococcus (Teniaceae) parasitic in humans and animals. Hydatid disease is distributed worldwide and is mainly prevalent in Europe, northern Africa, Asia, South America and Oceania. In my country, it is mainly found in Xinjiang, Ningxia, Qinghai, Tibet, Gansu, Aba and Ganzi in Sichuan, Diqing in Yunnan, northern Shaanxi and Inner Mongolia West etc. At present, there are 6 species of Echinococcus reported in the world, and 3 species are distributed in my country, namely E. granulosus, E. multilocularis and E. shiquicus). Echinococcus gra...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/20C07K16/18G01N33/577G01N33/558G01N33/58G01N33/543G01N33/53G01N33/52C12R1/91
CPCC07K16/18G01N33/577G01N33/558G01N33/587G01N33/54346G01N33/5308G01N33/52G01N2333/43543
Inventor 汪俊云石锋高春花杨玥涛危芙蓉张璟贾孝凯王颖
Owner 中国疾病预防控制中心寄生虫病预防控制所国家热带病研究中心
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