Detection method of aflatoxin M1

An aflatoxin and detection card technology, which is used in measuring devices, biological tests, material inspection products, etc., can solve the problem that aflatoxin detection technology cannot obtain safe "non-toxic" milk sources, and it is difficult to ensure the quality and safety of milk sources. Environmental pollution and other issues

Inactive Publication Date: 2019-08-30
SHENZHEN GRADUATE SCHOOL TSINGHUA UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0003] In view of the lack of fast, cheap, accurate and efficient aflatoxin and other residue detection technologies in current dairy products companies, and the lack of access to safe "non-toxic" milk sources, if a test kit for aflatoxin detection in milk can be formed , will help the detection of toxic aflatoxin residues in raw milk and its milk products, and solve the current...

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  • Detection method of aflatoxin M1
  • Detection method of aflatoxin M1
  • Detection method of aflatoxin M1

Examples

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Embodiment 1

[0067] Embodiment 1, the preparation of the antibody of the anti-aflatoxin M1 of fluorescence labeling

[0068] Anti-aflatoxin M1 mouse monoclonal antibody for test: product of Shenzhen Zhenkang Technology Co., Ltd., product number Ab012.

[0069] The first step is to reduce the disulfide bond in the anti-aflatoxin M1 mouse mAb with dithiothreitol to release the free thiol. Add 5 μL of DTT with a concentration of 1 M to 500 μL of anti-aflatoxin M1 mouse monoclonal antibody with a concentration of 2 mg / mL, incubate at room temperature for 30 min, and then purify with an ultrafiltration tube with a pore size of 30 kDa. Mix 10 mg of amino-functionalized CdSe / ZnS fluorescent quantum dots (Shenzhen Zhenkang Technology Co., Ltd., product number CQ003) with 0.01 mM sulfo-SMCC, incubate at room temperature for 30 min and desalt by ultrafiltration. The second step is to mix the anti-aflatoxin M1 mouse monoclonal antibody containing free thiol with desalted amino-functionalized CdSe / Zn...

Embodiment 2

[0070] Example 2, Preparation of Aflatoxin M1-Coupled BSA Antigen Labeled by Superparamagnetic Particles

[0071] 1. Preparation of superparamagnetic particles

[0072] Take 100mg of magnetic Fe with a particle size of 100-300 nanometers 3 o 4 Nanoparticles were treated with 0.1M HCl and washed. Then it was dispersed in 100ml of ethanol and water mixture (ethanol:water=1:4, V / V). Add 0.5 ml of concentrated ammonia water (concentration: 25%) under ultrasonic conditions. Afterwards, 1 ml of tetraethyl silicate was added and reacted for 24 hours. Disperse in 100ml ethanol after washing, add 0.5ml concentrated ammonia water (concentration is 25%), then add 1ml OTMS, react for 24 hours, disperse in chloroform after washing, then add 1gPMA-ODE (polymethacrylate-octadecyl Alkene), after drying, add ammonia water to dissolve, and obtain the surface carboxyl functionalized and coated SiO 2 superparamagnetic Fe 3 o 4 Particles, the diameter of which is 4-10 μm.

[0073] 2. Labe...

Embodiment 3

[0078] Embodiment 3, the preparation and use method of detecting aflatoxin M1 kit

[0079] 1. Preparation of kit for detecting aflatoxin M1

[0080] The present invention is used for detecting the kit of aflatoxin M1, comprises:

[0081] (1) Fluorescence-labeled anti-aflatoxin M1 antibody solution, the solvent is 0.02M PBS buffer at pH 7.4 containing 0.1% (0.1g / 100mL) BSA; the antibody concentration is 10 μg / mL). Specifically, the concentration of the 1 mg / mL fluorescently labeled anti-aflatoxin M1 antibody prepared in Example 1 was diluted 100 times with 0.02M PBS buffer solution of pH 7.4 containing 0.1% (0.1g / 100mL) BSA Subpackage.

[0082](2) Solution of aflatoxin M1 coupled with BSA antigen labeled with superparamagnetic particles: the solvent is 0.02M PBS buffer solution with pH 7.4; the concentration of superparamagnetic particles labeled with antigen is 100,000 / mL. Specifically, the aflatoxin M1-coupled BSA antigen labeled with superparamagnetic particles prepared i...

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Abstract

The invention discloses a method for detecting aflatoxin M1. The invention provides application of a kit to detection on the aflatoxin M1. The kit contains a fluorescently-labeled anti-aflatoxin M1 antibody, a superparamagnetic particle labeled aflatoxin M1 coupling protein antigen and a micro-fluidic chip detection card; the micro-fluidic chip detection card comprises a bottom plate and a micro-fluidic chip; the micro-fluidic chip is provided with a detection passage; the detection passage is formed by communicating a bottom passage with a sampling passage and a liquid suction passage which are respectively positioned at both ends of the bottom passage; openings of the sampling passage and the liquid suction passage respectively are a sampling hole and a liquid suction hole; the bottom passage is fixedly provided with a filter element; and the filter element is provided with a plurality of through holes for enabling sampling liquid to flow through. The detection kit and the detectionmethod, which are provided by the invention, are used for detecting the aflatoxin M1, and are high in detection sensitivity, short in detection time, low in detection cost and easy to operate and popularize.

Description

technical field [0001] The invention relates to the fields of biotechnology and food safety detection, in particular to a method for detecting aflatoxin M1. Background technique [0002] Aflatoxins (AFs) are a class of highly toxic mycotoxins widely present in feed and food storage, and are secondary metabolites of Aspergillus flavus and Aspergillus parasiticus. Because these mycotoxins are carcinogenic, hepatotoxic and mutagenic to animals and humans, they pose serious hazards to animal and human health. Among the several aflatoxins that have been found, aflatoxin B1 has the strongest toxicity, and it is widely found in moldy feed. After ingesting aflatoxin B1, dairy cows metabolize into aflatoxin M1 in vivo. Both aflatoxin B1 and M1 are classified as Class I carcinogens by the International Agency for Research on Cancer. [0003] In view of the lack of fast, cheap, accurate and efficient aflatoxin and other residue detection technologies in current dairy products compani...

Claims

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Application Information

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IPC IPC(8): G01N33/58G01N33/533G01N33/543
CPCG01N33/588G01N33/533G01N33/54326G01N33/54346
Inventor 马岚吴峰岑瑜毛茅
Owner SHENZHEN GRADUATE SCHOOL TSINGHUA UNIV
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