Immunofluorescent reagent used for detecting Muscovy duck gosling blast diseases

An immunofluorescence and goose plague technology, applied in fluorescence/phosphorescence, measuring devices, instruments, etc., can solve the problems of lack of rapid diagnostic reagents for gosling plague in Muscovy ducks, difficulties in clinical diagnosis, etc., and achieves innovative, practical, and accurate Highly specific and highly specific effects

Active Publication Date: 2012-04-25
INST OF ANIMAL HUSBANDRY & VETERINARY FUJIAN ACADEMY OF AGRI SCI
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  • Abstract
  • Description
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  • Application Information

AI Technical Summary

Problems solved by technology

Although the indirect immunofluorescence method established with gosling monoclonal antibody has been reported, there has not been a direct immunofluorescence diagnostic reagent mediated by monoclonal antib

Method used

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  • Immunofluorescent reagent used for detecting Muscovy duck gosling blast diseases
  • Immunofluorescent reagent used for detecting Muscovy duck gosling blast diseases
  • Immunofluorescent reagent used for detecting Muscovy duck gosling blast diseases

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Experimental program
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Effect test

preparation example Construction

[0020] 3.1 Preparation of GPV-infected cells: Muscovy duck gosling plague virus (GPV) was inoculated on a 96-well MDEF cell culture plate that had grown into a single layer, and normal MDEF cells were set as negative controls; cultured at 37°C and 5% CO2 When the cell lesion reaches about 30%, discard the culture supernatant, wash once with sterile PBS or normal saline, add 100 μL of cold methanol to each well, fix at 4°C for 30 min, discard the fixative and wash once at -40 ℃ frozen for later use.

[0021] 3.2 Preparation of tissue frozen sections: take the liver, spleen and pancreas of Muscovy duck artificially infected with Muscovy duck gosling plague virus (GPV) or the embryo heart of dead Muscovy duck inoculated with GPV, and cut into 0.5cm 3 About the size, placed on the tissue fixer of the cryostat, cut into 6-8um thin slices after the tissue was frozen, adhered to the glass slide, fixed in cold acetone for 10-15 minutes, and frozen at -20°C for later use.

[0022] 3.3...

experiment example

[0036] According to the operation method of direct immunofluorescence assay (DFA), after appropriately diluting the fluorescent antibody, take GPV, MPV, MDRV, ARV, PMV and other infected cells and normal control cells respectively, add 1:150 diluted fluorescent reagent 30ul to each well, and carry out DFA test. The results show that weak positive fluorescence can be seen in the nucleus of MDEF cells infected with GPV at the initial stage, and strong bright green fluorescence appears in the nucleus 36 hours after infection, and then the fluorescence gradually appears in the whole cell and goes out with the disintegration of the cell. Diffusion; while other virus-infected cells and normal MDEF cells did not show fluorescence. It shows that GPV immunofluorescence reagent can be used for the identification and antigen localization of GPV-infected cells.

[0037] Table 2. Identification of GPV antigens by immunofluorescence assay (DFA)

[0038]

[0039] Note: Judgment criteria...

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Abstract

The invention discloses an immunofluorescent reagent used for detecting muscovy duck gosling blast diseases, which is characterized in that an anti-GPV hybridoma cell strain D11 strain is used for preparing anti-GPV monoclonal antibody, then the GPV monoclonal antibody is used for labeling fluorescein isothiocyanate (FITC) to prepare the immunofluorescent reagent used for detecting muscovy duck gosling blast diseases; the anti-GPV hybridoma cell strain D11 strain is preserved in CCTCC, the preservation date is April 15th, 2011, and the preservation number is C201120. The invention also discloses an application of the immunofluorescent reagent used for detecting muscovy duck gosling blast diseases in clinical quick diagnosis of the muscovy duck gosling blast diseases, and an application ofpositioning and identifying of goose parvovirus antigen in tissue cells.

Description

technical field [0001] The invention relates to an immunofluorescent reagent for detecting the plague of muscovy duck and goose. Background technique [0002] Muscovy duck goose plague is caused by goose parvovirus (Goose parvovirus, GPV), a viral infectious disease characterized by diarrhea, intestinal mucosa shedding and embolism in young muscovy ducks under one month old. The disease has been in Fujian Province since 1997 Muscovy duck breeding areas in Putian, Fuqing and other places broke out. At present, it has occurred in various duck breeding areas across the country, with a morbidity rate of 50%-70% and a fatality rate of 40%-65%. Clinically, the disease is not only often co-infected with Muscovy duck parvovirus disease (three-week disease), but also often misdiagnosed as Muscovy duck parvovirus disease by grassroots veterinarians. Since the hyperimmune serum against gosling plague has a good therapeutic effect on the disease, rapid diagnosis is the premise of effec...

Claims

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Application Information

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IPC IPC(8): G01N33/577G01N21/64
Inventor 陈少莺朱小丽陈仕龙程晓霞林锋强王燕玲王劭李兆龙
Owner INST OF ANIMAL HUSBANDRY & VETERINARY FUJIAN ACADEMY OF AGRI SCI
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