Colloidal gold test card for quickly testing ergot alkaloid and preparation method thereof

A technology of ergot alkaloids and detection cards, which is applied in the direction of measuring devices, instruments, scientific instruments, etc., can solve the problems of limited application range, high detection cost, cumbersome operation, etc., and achieve high detection accuracy, strong specificity, portable convenient effect

Inactive Publication Date: 2012-10-03
南通市产品质量监督检验所 +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In general, the quantitative determination of alkaloid content by HPLC has good sensitivity and specificity, but the operation is cumbersome and time-consuming, and it is not suitable for the detection of large quantities of samples, and requires expensive instruments and equipment, and the detection cost is high , and require operators to have certain professional knowledge to operate, so the scope of application is limited; colloidal gold immunochromatography detection technology is a rapid immunodiagnostic technology, using colloidal gold as a marker, within a short time (within 10 min) Intuitive results can be obtained, it does not need to separate unreacted conjugates from reacted conjugates, saves tedious sample addition and washing steps, the operation process is simple and fast, the specificity and sensitivity are also high, and the detection cost is low. Compared with other methods, it can realize the real one-step rapid detection, and non-professionals can also operate it, which is in line with the development direction of modern analysis and detection technology

Method used

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  • Colloidal gold test card for quickly testing ergot alkaloid and preparation method thereof
  • Colloidal gold test card for quickly testing ergot alkaloid and preparation method thereof
  • Colloidal gold test card for quickly testing ergot alkaloid and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0020] Example 1: Preparation of Complete Antigen

[0021] Preparation of lysergic acid-keyhole limpet hemocyanin (lysergic acid-KLH) and lysergic acid-ovalbumin (lysergic acid-OVA): ① Dissolve 75~85 mg lysergic acid in 10 mL of hot dioxane, then add into 100-120 mg carbodiimide (EDC) aqueous solution (5 mL). ② Dissolve 20-40 mg keyhole limpet hemocyanin (KLH) or ovalbumin (OVA) in 10 mL PBS, and add dropwise to the mixture prepared in the previous step. ③After stirring at room temperature for 10 hours, store at 4°C overnight. Then dialyzed with PBS for 2 days to obtain lysergic acid-keyhole limpet hemocyanin (lysergic acid-KLH) and lysergic acid-ovalbumin (lysergic acid-OVA) respectively, which were freeze-dried and stored at -20°C for later use.

Embodiment 2

[0022] Example 2: Preparation and purification of anti-ergot alkaloid polyclonal antibody

[0023] Three healthy male New Zealand white rabbits were immunized with the immunizing antigen (lysergic acid-KLH) obtained in Example 1. One week before immunization, blood was collected from the rabbit's ear vein as a negative control. Weigh 1 mg of lysergic acid-KLH, dissolve it with 2 mL of 0.1 mol / L PBS (sterilized by autoclaving), then take 1 mL of the dissolved antigen and mix it with 1 mL of complete Freund’s adjuvant. Point subcutaneous injection. One month later, the first booster immunization was carried out by intramuscular injection of the limbs, the dose was the same as that of the first immunization, mixed with the same amount of incomplete Freund's adjuvant and fully emulsified. A total of 5 times of immunization, each interval of 2 weeks. Before each booster immunization, blood was collected from the rabbit's ear vein, and the titer of antiserum was determined by ind...

Embodiment 3

[0024] Embodiment 3: the preparation of colloidal gold solution

[0025] Prepare 100 mL of 0.01% chloroauric acid solution with fresh double-distilled water, place it in a conical flask, heat it to boiling (about 110°C) with a constant temperature electromagnetic stirrer, and quickly add the 4.5 mL of 1% trisodium citrate solution kept at ℃, keep the temperature and rotational speed constant, continue to stir and heat for about 15 min, until the solution turns clear wine red; cool at room temperature, and store in the refrigerator at 4°C for later use.

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Abstract

The invention discloses a colloidal gold test card for quickly testing ergot alkaloid and a preparation method thereof. A test paper strip is packaged in a shell of the test card; a sample adding hole and an observation hole are formed on the shell of the test card; the test paper strip comprises a base plate; a piece of water-absorbing paper, a nitrocellulose membrane, a glass fiber membrane and a sample pad are arranged on the base plate in turn; the glass fiber membrane is soaked with an ergot alkaloid antibody marked by colloidal gold; and two lines of envelope antigen ergotic acid-OVA and goat anti-rabbit antibody are sprayed on the nitrocellulose membrane. The colloidal gold test card provided by the invention is conveniently taken and conveniently operated; the colloidal gold test card is used for on-site semi-quantitatively testing raw grains and the content of the ergot alkaloid in the product of the raw grains; the testing accuracy is high and the specificity is strong; the test for the ergot alkaloid is no longer limited by professionals in a professional institute; and all livestock farms, grain purchasing stations, all-level check quarantine inspection departments and consumers can test in real time.

Description

technical field [0001] The invention relates to a preparation method of a colloidal gold detection card for rapidly detecting ergot alkaloids. Background technique [0002] Ergot is a purple-black, long-horned sclerotia with a fungal structure formed by ergot bacteria in the ovary of parasitic plants. It is shaped like an animal horn, so it is called ergot. Most of its hosts are grasses. Wheat, corn, barley, rice, millet, oats, sorghum, and rye grown all over the world can be infected by ergot. Ergot poisoning is caused by a series of active alkaloids in grains infected with ergot fungi. The toxic components of ergot toxins are mainly a series of alkaloid derivatives based on lysergic acid, such as ergotoxin, ergotamine, ergonovine, ergosine, Ergocristine (ergocristine), etc. Ergot alkaloids are a general term for a class of substances that are commonly used in medicine to contract the uterus, promote labor, treat migraines, and high blood pressure. On the other hand, e...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/543
Inventor 陈刚周鸣镝张东升黄伟东邵彪龚燕杨婷婷匡群张海涛
Owner 南通市产品质量监督检验所
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