Tobacco cucumber mosaic virus field rapid detection test paper preparation method

A technology for detecting cucumber mosaic virus and test paper, which can be applied to measuring devices, instruments, scientific instruments and other directions, and can solve the problems of low titer of antiserum and unsatisfactory detection effect.

Inactive Publication Date: 2008-07-02
HENAN AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] However, both of these methods require the extraction and purification of viruses or nucleic acids in the samples, and equipment such as ultra-high-speed centrifuges and electron microscopes are require

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0013] Antibody preparation: To prepare tobacco-cucumber mosaic virus rabbit polyclonal antibody: Dilute the antigen, ie, purified CMV, to 2mg / ml with sterile saline, and then add Freund's adjuvant. The first time is a complete adjuvant, and the subsequent use is incomplete Adjuvant. The amount of antigen injection is 0.1~0.2mg / kg, 2~3Kg rabbits are selected, and the intravenous injection is done in three times. The amount of antigen injection is 0.03~0.1mg each time, and the interval is 10d. Bloodletting was performed 10 days after the last injection. Animals were fasted for 12 hours before bleeding, and 40-60ml of blood was used for ear vein bleeding. After collection, the blood cells were removed by centrifugation and stored at minus 20°C after purification. Goat anti-rabbit antibodies were also prepared.

[0014] Preparation of colloidal gold particles: Take 0.6 ml of 1% chloroauric acid in 30 ml of ultrapure water and boil it. Quickly add 0.9ml of 1% sodium citrate pre-warmed...

Embodiment 2

[0019] Example 2: In this example, when preparing colloidal gold particles: 0.6 ml of 1% chloroauric acid was heated and boiled in 30 ml of ultrapure water. Quickly add 0.9ml of 1% sodium citrate pre-warmed at 37°C at a time, the solution gradually changes from blue to purple-red, boil for 3min, and add water to 30ml. After cooling to room temperature, the pH was adjusted to 7.5 with 0.2mol / L potassium carbonate. Others are the same as in Example 1.

Embodiment 3

[0020] Example 3: In this example, when preparing colloidal gold particles: 0.6 ml of 1% chloroauric acid was heated and boiled in 30 ml of ultrapure water. Quickly add 0.9ml of 1% sodium citrate pre-warmed at 37°C at a time, the solution will gradually change from blue to purple-red, boil for 4min, and add water to 30ml. After cooling to room temperature, the pH was adjusted to 7.4 with 0.2mol / L potassium carbonate. Others are the same as in Example 1.

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PUM

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Abstract

The invention discloses a preparation method of test strip for rapid field detection of tobacco cucumber mosaic virus, which comprises the following steps of: preparing an antibody, preparing colloidal gold, preparing a colloidal gold-antibody conjugate, preparing a nitrocellulose filter, and preparing a colloidal gold chromatographic test strip. The prepared test strip for rapid field detection of tobacco cucumber mosaic virus according to the technical proposal has an optimum detection temperature of 15-35DEG C, sample detection amount of 0.1-0.3g and tobacco sample area of 3-5cm<2>. The detection method comprises charging tobacco leaf in a plastic bag containing buffer, grinding, immersing the test strip in the buffer, reacting for 30min, and observing the result. The false positive rate is less than 1%. The invention has the advantages of convenient application and high reliability; and is especially suitable for rapid field detection.

Description

Technical field [0001] The invention belongs to detection materials, and particularly relates to a method for preparing a test strip for rapid field detection of tobacco-cucumber mosaic virus. Background technique [0002] Tobacco and cucumber mosaic disease occurs in all tobacco-producing areas in my country. In recent years, the damage has gradually become heavier than ordinary mosaic disease. It causes large economic losses every year in Henan, Shandong, Shaanxi, Sichuan, Heilongjiang, Liaoning and other provinces. One of the important viral diseases in our country. The nucleic acid of Cucumber mosaic virus (CMV) is a triad of linear positive-strand ssRNA, which is mainly overwintered on overwintering vegetables and perennial weeds. In the following spring, toxic winged aphids stabbing smoking leaves to form initial infections , The field is infested by aphids and friction contact. In the early stage of tobacco infection, only symptoms such as heart leaf veins, mosaic and othe...

Claims

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Application Information

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IPC IPC(8): G01N33/544G01N33/532
Inventor 高玉千邱立友梁振普戚元成
Owner HENAN AGRICULTURAL UNIVERSITY
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