A method for detecting lily mottle virus with monoclonal antibody gold label test
A lily mottle virus and lily technology, which is applied in the field of monoclonal antibody gold label test to detect lily mottle virus, can solve unused problems, achieve high accuracy, strong pertinence, and reduce economic losses
Active Publication Date: 2016-10-05
NORTHWEST INST OF ECO ENVIRONMENT & RESOURCES CAS
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But so far there is no relevant report about the preparation of LMoV monoclonal antibody at home and abroad, and the monoclonal antibody of LMoV has not been used in colloidal gold immunochromatography test
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[0026] 1. Preparation of natural LMoV antigen
[0027] The oriental lily leaves that were naturally infected with LMoV and showed mottled symptoms were collected in Lanzhou, Gansu, and were verified as LMoV single infection by RT-PCR. The method was slightly modified by referring to Chen Jianping’s method, and 0.05 mol / L Tris-HCl (pH 7.4, Contains 0.01 mol / L EDTA.Na 2 , 0.01 mol / L MgSO 4 , 0.1 mol / L NaCl, 0.1% (w / v) PVPP and 0.1% (w / v) β-mercaptoethanol) as the buffer to extract LMoV. The purified virus was negatively stained with 2% phosphotungstic acid (pH 6.7) and placed under a JEM-1230 transmission electron microscope from Japan JEOL Company to observe the particle morphology; it was verified as LMoV by RT-PCR and Western blotting.
[0028] 2. Preparation of monoclonal antibodies
[0029] After fully emulsifying the purified natural LMoV antigen with the same amount of Freund's complete adjuvant, immunize 8-week-old BALB / C mice, 0.3 mL each; three weeks later, switch t...
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Abstract
The invention provides a method for detecting the lily mottle virus (LMoV) by using monoclonal antibodies through colloidal gold immunoassay. The method comprises the following steps: extracting and purifying natural LMoV from oriental lily leaves naturally infected with LMoV and having obvious mottle symptoms; preparing the monoclonal antibodies Mab1 and Mab2 from purified LMoV particles; labeling the first monoclonal antibody Mab1 with colloidal gold and using the labeled first monoclonal antibody Mab1 as a missing marker; and allowing a to-be-detected solution to specifically bind with the second monoclonal antibody Mab2 and a goat anti-mouse IgG purified antibody at a detection line and a contrast line so as to realize color development and determining negative and positive results according color development situations of the detection line and the contrast line. The method does not need special training and can rapidly, easily and accurately detect the lily mottle virus.
Description
technical field [0001] The invention relates to a method for detecting lily mottle virus by monoclonal antibody gold standard test, in particular to a method for detecting lily mottle virus by applying monoclonal antibody to colloidal gold immunochromatography test. Background technique [0002] Lily mottle virus (LMoV) is the main virus that seriously endangers the production of lily. It is widely distributed in the world, especially in all temperate regions suitable for the growth of its hosts and regions rich in aphid vectors. LMoV infection of Liliaceae plants often causes mottled stripes or even necrotic spots on the leaves, and later develops into flowers, curled and deformed leaves, and broken colored flowers, often accompanied by short plants, reduced flower and bulb yields, etc. LMoV belongs to the Potatoviridae, Potatovirus genus ( Potyvirus ), the virus particles are curved and linear and helically symmetrical, with a size of (650-900) nm × (11-15) nm. The viral...
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IPC IPC(8): G01N33/569G01N33/577G01N33/558
CPCG01N33/558G01N33/56983G01N33/577
Inventor 张玉宝陈延王亚军刘维谢忠奎王若愚郭志鸿王乐
Owner NORTHWEST INST OF ECO ENVIRONMENT & RESOURCES CAS