Preparation and detection method of aflatoxin G1 enzyme-linked immunosorbent assay kit

An enzyme-linked immunosorbent reagent and aflatoxin technology, which is used in measuring devices, instruments, scientific instruments, etc., can solve the problems of complex operation, high false positive rate, and large pollution, and achieves convenient use, simple kit and high sensitivity. Effect

Inactive Publication Date: 2014-05-14
JIANGSU WISE SCI & TECH DEV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

These detection methods have played a good role in different periods, but due to more or less defects such as complicated operation, heavy pollution, high requirements for equipment, high false positive rate, and long detection time, it is difficult to adapt to the requirements of today's society.

Method used

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  • Preparation and detection method of aflatoxin G1 enzyme-linked immunosorbent assay kit
  • Preparation and detection method of aflatoxin G1 enzyme-linked immunosorbent assay kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0021] The pretreatment of embodiment 1 experimental material and sample

[0022] Preparation of solid-phase antigen on coated plate:

[0023] Add AFG1-BSA with 50 mmol / L Na 2 C0 3 -NaHC0 3 Dilute the pH9.6 buffer solution to 10 mg / L coating solution, add 100 μL to each well of a 96 (or 48)-well microplate, incubate at 37 °C in the dark for 2 h, discard the coating solution, wash twice, Add 150 μL of the above Na containing 3% BSA 2 CO 3 -NaHCO 3 Block with buffer, incubate at 37°C in the dark for 1.5 h, discard the blocking solution, dry in the air, seal the strips and store in a freezer at -20°C.

[0024] Preparation of reagents:

[0025] (1) Standard aflatoxin G1: (0 ng / mL, 0.025 ng / mL, 0.1 ng / mL, 0.25 ng / mL, 0.5 ng / mL, 1.5 ng / mL), diluted from pure AFG1, Diluent is methanol: water volume ratio is 4:6;

[0026] (2) Horseradish peroxidase-labeled goat anti-rabbit antibody freeze-dried product (HRP—goat anti-rabbit antibody);

[0027] (3) Washing solution: 68.8 g...

Embodiment 2

[0046] Example 2 Method determination

[0047] Precautions before measurement

[0048] 1. Return all reagents and microplates to room temperature before use;

[0049] 2. Immediately after use, put all the reagents back to 2~8 ℃;

[0050] 3. Do not let the micropores dry during use;

[0051] 4. The repeatability in ELISA analysis largely depends on the consistency of plate washing, and the correct plate washing operation is the key point in the ELISA assay procedure;

[0052] 5. During all constant temperature incubations, avoid light exposure and cover the microwell plate with a cover film;

[0053] 6. Take out the required number of microporous plates and frames, put the unused microporous plates into the original tin foil bag and reseal it together with the provided desiccant, and store at 2-8 ℃.

[0054] Specific measurement steps

[0055] 1. Take the required reagents and microwell plates out of the refrigerated environment, equilibrate at room temperature for 30 mi...

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Abstract

The invention discloses an enzyme-linked immunosorbent assay kit for detecting aflatoxin G1 in peanuts, cereals and milk products. In the kit, based on a competitive ELISA (enzyme-linked immunosorbent assay) method, ELISA plate micropores are coated with the aflatoxin G1 antigen. The detection comprises the following steps: adding a standard product or sample solution, an aflatoxin G1 antibody and an enzyme marker; competitively combining the coating antigen and the aflatoxin G1 added into the sample with the aflatoxin G1 antibody; washing off the uncombined antigen and antibody; further combining the combined antigen-antibody compound with the enzyme marker; washing off the uncombined antigen and antibody, and developing color through a TMB substrate; adding a reaction stop solution, and detecting with a 450 / 630nm dual-wavelength microplate reader, wherein the aflatoxin G1 concentration in the sample is inversely proportional to the absorption light intensity; and comparing with the standard curve and multiplying with corresponding dilution ratio to obtain the content of aflatoxin G1 in the sample.

Description

technical field [0001] An enzyme-linked immunoassay kit and a detection method for detecting aflatoxin G1 belong to the technical field of biological and food safety detection, and can be used for detecting the content of aflatoxin G1 (AFG1) in peanuts, grains, and milk products. Background technique [0002] Aflatoxin is mainly a secondary metabolite secreted by Aspergillus flavus and Aspergillus parasiticus, and is a natural toxic compound that can cause various damages to humans and animals. More than 20 types of aflatoxin (AFT) have been discovered, among which aflatoxin B1 is the most toxic and carcinogenic substance. The toxicity of aflatoxin G1 is second only to aflatoxin B1, and it is a strong carcinogen. Studies have shown that it has a stronger potential to induce kidney tumors than AFB1. It is one of the main contaminating mycotoxins in the diet of residents in areas with high incidence of esophageal cancer and lung cancer in the south of Taihang Mountains in my...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/577
CPCG01N33/543G01N33/54373G01N2333/38
Inventor 杜道林王永美
Owner JIANGSU WISE SCI & TECH DEV
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