Colloidal gold test strip capable of simultaneously detecting waterfowl source parvoviruses and preparation method

A technology of colloidal gold test paper and parvovirus, applied in the field of veterinary medicine, can solve the problems of limited application and narrow use range, and achieve the effects of reducing false positives, high sensitivity, and improving specificity and sensitivity

Inactive Publication Date: 2017-05-10
INST OF ANIMAL HUSBANDRY & VETERINARY FUJIAN ACADEMY OF AGRI SCI +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Colloidal gold test strips in the prior art can only detect a kind of waterfowl-deriv

Method used

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  • Colloidal gold test strip capable of simultaneously detecting waterfowl source parvoviruses and preparation method
  • Colloidal gold test strip capable of simultaneously detecting waterfowl source parvoviruses and preparation method
  • Colloidal gold test strip capable of simultaneously detecting waterfowl source parvoviruses and preparation method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] Example 1: Preparation of Gosling Plague Virus Monoclonal Antibody

[0027] 1.1 Immunization of mice

[0028] Concentrated and purified gosling plague virus was used as antigen, emulsified with Freund's complete adjuvant, injected subcutaneously into mice at multiple points, and boosted with the same antigen plus Freund's incomplete adjuvant for 5 times every 2 weeks. One week after the last immunization, blood was collected to prepare serum, and the titer was determined by indirect ELISA method. Mice whose serum titer reached 1:1000 and above were given a booster immunization without adjuvant antigen, and cell fusion was carried out 3 days later.

[0029] 1.2 Preparation of myeloma cells

[0030] 48 hours before fusion, SP2 / 0 myeloma cells were expanded and cultured in cell culture dishes with a diameter of 9 cm, and 10 mL of DMEM medium (product of Gibco Company) containing 10% fetal bovine serum was added to each culture dish, and placed at 37°C, 5%CO 2 Culture in...

Embodiment 2

[0053] Example 2: Colloidal gold labeling of monoclonal antibody 3D9

[0054] 2.1 Preparation of colloidal gold

[0055] Take 1mL of 10g / L chloroauric acid solution and add it to the Erlenmeyer flask that has been treated with soaking acid and siliconization, add 100mL deionized water, seal it with tin foil, and put it in the microwave oven. Bring the solution to boil on high heat for 3 minutes, take out the Erlenmeyer flask, quickly add 1.5mL of 10g / L trisodium citrate solution, and mix well at the same time. The solution will change from pale yellow to colorless to grayish black. Seal with tin foil, put the Erlenmeyer flask into the microwave oven, heat it on medium-high heat for 3 minutes, after the solution turns wine red, heat it for another 3 minutes, after it cools down to room temperature, fill it up to 100mL with deionized water, and filter it with a 450nm filter membrane , stored at 4°C protected from light.

[0056] 2.2 Colloidal gold labeling

[0057] Centrifug...

Embodiment 3

[0058] Embodiment 3: the assembly of colloidal gold test strip

[0059] 3.1 Preparation of gold standard pad

[0060] Soak the glass cellulose membrane in the colloidal gold-labeled monoclonal antibody 3D9, let it stand at 37°C for 30 minutes, and dry it at room temperature to form a gold label pad, which is stored at 4°C.

[0061] 3.2 Preparation of nitrocellulose membrane (NC membrane)

[0062] Spray 2 mg / mL of monoclonal antibody 5F4 on the test line (T) on the NC membrane with a membrane spraying system; spray 1 mg / mL of goat anti-mouse IgG antibody on the quality control line (C) on the NC membrane with a membrane spraying system ). Dry at room temperature, soak in 20g / L BSA solution, let stand at room temperature for 30min, wash with PBST 3 times, 5min each time, dry at room temperature, that is, NC membrane, and store at 4°C.

[0063] 3.3 Assembly

[0064] Paste the sample pad, the prepared gold standard pad, NC film and absorbent paper on the plastic bottom plate i...

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Abstract

The invention relates to the field of animal medicine, and discloses a colloidal gold test strip capable of simultaneously detecting waterfowl source parvoviruses and a preparation method. The colloidal gold test strip comprises a base plate, a sample pad, a gold mark pad, a nitrocellulose membrane and an absorbent pad, the gold mark pad is coated with colloidal gold mark protein with gosling plague virus monoclonal antibodies I, detection lines and quality control lines are sequentially arranged on the nitrocellulose membrane along flowing directions of samples, the detection lines of the nitrocellulose membrane are coated with gosling plague virus monoclonal antibodies II, and the quality control lines of the nitrocellulose membrane are coated with goat-anti-mouse IgG (intravenous gamma globulin) antibodies. The colloidal gold test strip can simultaneously detect goose parvoviruses, Muscovy duck parvoviruses and novel Muscovy duck parvoviruses, false positive of reaction is reduced to a great extent, and specificity and sensitivity of detection are improved.

Description

technical field [0001] The invention relates to the field of veterinary medicine, in particular to a colloidal gold test strip capable of simultaneously detecting waterfowl-derived parvoviruses and a preparation method thereof. Background technique [0002] Waterfowl-derived parvovirus refers to the parvovirus that infects waterfowl (here mainly refers to domesticated ducks and geese, wild geese, swans, etc.) virus etc. [0003] The detection methods of parvovirus in waterfowl include virus isolation and identification, molecular biology detection and serological detection. The virus isolation and identification method is to culture the disease material in specific cells, and after several passages, it is judged according to whether specific lesions appear, and then according to the observation of electron microscope morphology, which takes a long time, requires a lot of equipment, and requires high technical level of personnel . PCR technology, loop-mediated isothermal a...

Claims

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Application Information

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IPC IPC(8): G01N33/577G01N33/569G01N33/558G01N33/531
CPCG01N33/531G01N33/558G01N33/56983G01N33/577G01N2333/015
Inventor 程龙飞张长弓黄瑜何琼贾志娟陈慧敏傅秋玲傅光华施少华万春和刘荣昌陈红梅
Owner INST OF ANIMAL HUSBANDRY & VETERINARY FUJIAN ACADEMY OF AGRI SCI
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