PVX Yunnan isolate TAS-ELISA test kit and its preparing method

A detection kit and the technology of the kit are applied in the field of enzymology or biological devices, and can solve the problems of no TAS-ELISA detection kit, no report on the large-scale detection of plant viruses, etc.

Inactive Publication Date: 2005-05-04
YUNNAN ACAD OF AGRI SCI +2
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Problems solved by technology

[0007] So far, reports on the study of plant viruses by TAS-ELISA method have increased in recent years, but the large-scale detection of p

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  • PVX Yunnan isolate TAS-ELISA test kit and its preparing method
  • PVX Yunnan isolate TAS-ELISA test kit and its preparing method

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Embodiment Construction

[0060] The collection and detection process is as follows: the diseased plant samples are collected in the field, and detected by electron microscope and standard antibody to contain a large amount of PVX, and only contain the virus. The extraction buffer was 0.2M PBS (pH7.4, containing 0.5% mercaptoethanol, 0.01M EDTA).

[0061] The purification process is:

[0062] Cut 50g of diseased leaves into pieces, add 100mL of extraction buffer, fully homogenize with an electric mixer, filter with double gauze, add 30% chloroform to the filtrate, stir at 4°C for 30min, centrifuge at 3000g for 10min, and collect the supernatant;

[0063] Add 8% PEG, 0.1M NaCl to the supernatant, dissolve and place at 4°C for 4h or overnight; centrifuge at 10000rpm, 4°C, 20min, take the precipitate, suspend it with 0.05M citric acid buffer (pH7.4), centrifuge at 10000rpm, 4°C, 10min, a precipitate was obtained;

[0064] Suspend the precipitate with 0.01M PBS (pH7.4), centrifuge at 10000rpm, 4°C, 10min...

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Abstract

This invention relates to a kind of potato virus X (PVX) yunnan outlier three antibody sandwich enzyme immune adsorbing rule (TAS-ELISA) detection kit and its preparation method. The kit comprises positive and negative contrast, PVX multi-clone antibody, PVX single clone antibody, enzyme standard antibody and other materials and medicines. The PVX multi-clone antibody is got from rabbit immune adsorbing blood serum after separately purifying the PVX yunnan outlier. The PVX single clone antibody is got from mouse immune, cell fusion, clone and purification after separately purifying the PVX yunnan outlier. The detection sensitivity of the TAS-ELISA detection kit in this invention can reach 0.01ng/ml. It coincides with the comparing accuracy rate of electrical glass detection result, and it is 100 times than indirect ELISA and DAS-ELISA detection sensitivity.

Description

Technical field: [0001] The invention relates to a triple-antibody sandwich enzyme-linked immunosorbent assay (TAS-ELISA) detection kit for potato virus X (PVX) Yunnan isolate and a preparation method thereof, belonging to the field of enzymology or biological devices. Background technique: [0002] Plant viruses are important pathogens of crop virus diseases. With the development of plant virus-free seedling industrialization and the rise of modern facility agriculture, the detection and prevention and control technology of plant viruses has been paid more and more attention. At present, the detection technology of plant virus has four methods: serological technology, indicator plant determination, electron microscope detection, and molecular detection (including the detection of viral nucleic acid and protein). Various methods confirm each other, and at the same time, there are limitations in detection sensitivity, specialization, equipment and facility conditions, and cos...

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Application Information

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IPC IPC(8): G01N1/34G01N33/53G01N33/531
Inventor 张仲凯周雪平杨录明汪继玲丁铭方琦
Owner YUNNAN ACAD OF AGRI SCI
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