Chemiluminescence detection kit for aflatoxins M1 and preparation method of chemiluminescence detection kit

A chemiluminescence detection, aflatoxin technology, applied in chemiluminescence/bioluminescence, analysis by chemical reaction of materials, etc., can solve the problems of long detection time, cumbersome time-consuming, weak stability, etc., to achieve rapid detection Convenience, simple and convenient operation, stable luminous value

Inactive Publication Date: 2018-02-13
太原瑞盛生物科技有限公司
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  • Abstract
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  • Claims
  • Application Information

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Problems solved by technology

Among them, high-performance liquid chromatography, gas chromatography-mass spectrometry, and liquid chromatography-mass spectrometry / mass spectrometry require large-scale precision instruments, which are cumbersome and time-consuming, with high detection costs, limited detection samples, and are not suitable for the detection of large batches of samples; CN 103513035 A (2014.01) A method to detect aflatoxin M 1 Test strip and method, the method adopts colloidal gold labeling aflatoxin M 1 The detection of antibodies and colloidal gold is characterized by the color visible to the naked eye, which has the disadvantages of large error, cumbersome operation, many processes, low sensitivity, and low precision; CN 104569381 A (2015.04) discloses a method for detecting yellow Aspergillus M 1 The method and the enzyme-linked immunosorbent assay kit, the kit adopts the enzyme-linked immunosorbent method for detection, and the enzyme-linked immunoassay method has the disadvantages of low sensitivity, difficulty in realizing full automation, and long detection time; CN 103091494 A (2013.05) discloses a aflatoxin M 1 The chemiluminescent enzyme-linked immunoassay kit and its use method, the kit uses horseradish peroxidase as a marker, and the disadvantage of using horseradish peroxidase for labeling is that the enzymatic process is greatly affected by the environment: such as disinfectant, Both pH and ions will affect it, and the stability is weak, thus affecting the measurement results

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] Embodiment 1: The formation and preparation of kit 1

[0027] 1. Kit 1 set-up

[0028] Aflatoxin M 1 The chemiluminescence detection kit, which contains the following components:

[0029] acridinium ester labeled aflatoxin M 1 Antibody;

[0030] Aflatoxin M coupled to carboxyl magnetic beads 1 antigen;

[0031] Chemiluminescence pre-excitation solution A and chemiluminescence excitation solution B;

[0032] Aflatoxin M 1 A series of calibrator solutions, the concentrations are: 0 ng / mL, 0.01 ng / mL, 0.05 ng / mL, 0.25 ng / mL, 1.25 ng / mL, 6.25 ng / mL;

[0033]The cleaning solution is specifically a Tris-HCl solution with a pH of 7.2 and a concentration of 25 mmol / L, which contains NaCl and 0.05% Tween-20 at a concentration of 0.15 mol / L.

[0034] 2. Conjugated with aflatoxin M 1 Preparation of Magnetic Microparticle Suspension of Antigen

[0035] (1) Take 1 mg of carboxyl magnetic particles in a 0.5 mL centrifuge tube, add 200 μL of 0.1 mol / L MES (pH 5.0) buffer, vo...

Embodiment 2

[0050] Embodiment 2: The formation and preparation of kit 2

[0051] 1. Assembly of Kit 2

[0052] Aflatoxin M 1 The chemiluminescence detection kit, which contains the following components:

[0053] acridinium ester labeled aflatoxin M 1 antigen;

[0054] Carboxyl magnetic beads coupled with aflatoxin M 1 Antibody;

[0055] Chemiluminescence pre-excitation solution A and chemiluminescence excitation solution B;

[0056] Aflatoxin M 1 A series of calibrator solutions, the concentrations are: 0 ng / mL, 0.01 ng / mL, 0.05 ng / mL, 0.25 ng / mL, 1.25 ng / mL, 6.25 ng / mL;

[0057] The cleaning solution is specifically a Tris-HCl solution with a pH of 7.2 and a concentration of 25 mmol / L, which contains NaCl and 0.05% Tween-20 at a concentration of 0.15 mol / L.

[0058] 2. Conjugated with aflatoxin M 1 Preparation of Magnetic Microparticle Suspension of Antibody

[0059] (1) Take 1 mg of carboxyl magnetic particles in a 0.5 mL centrifuge tube, add 200 μL of 0.1 mol / L MES (pH 6.0) b...

Embodiment 3

[0074] Embodiment 3: Aflatoxin M in actual samples 1 detection

[0075] Aflatoxin M of the present invention 1 The procedure for using the quantitative detection kit is as follows:

[0076] 1. Sample pretreatment

[0077] (1) Milk sample processing

[0078]Obtaining the milk test sample solution: Take 150 μL of fresh milk in a 500 μL centrifuge tube, centrifuge at 4°C for 10 min (3000 r / min), and discard the upper layer of fat. Pipette 25 μL of the centrifuged milk sample into a clean glass test tube, add 475 μL of 0.02 mol / L phosphate buffer for dilution.

[0079] (2) Milk powder sample processing

[0080] Take 1 g of sample in a 50 mL centrifuge tube, add 10 mL of 2% NaCl and 0.2 mol / L HCl-methanol mixture, and shake for 1 min. Centrifuge at 5000 r / min for 8 min, take 0.5 mL supernatant, add 0.05 mL, 0.5 mol / L NaOH solution, then add 9.45 mL 0.02 mol / L phosphate buffer, and mix well.

[0081] 2. Detection of the kit

[0082] (1) Add 100 μL of the sample to be tested,...

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Abstract

The invention discloses a chemiluminescence detection kit for aflatoxins M1 and a preparation method of the chemiluminescence detection kit. The chemiluminescence detection kit comprises an acridiniumester marker, magnetic particles coupled with an antigen or antibody, a calibrator solution, a cleanout fluid, a chemiluminescence pre-excitation liquid A and a chemiluminescence excitation liquid B.According to the chemiluminescence detection kit for aflatoxins M1, the magnetic separation chemiluminescence technology is taken as a detection means, and the acridinium ester marking technology isused together. The chemiluminescence detection kit for aflatoxins M1 is simple and convenient to operate, mild in reaction condition and stable in lighting value and is less influenced by external conditions. Compared with the prior art, the chemiluminescence detection kit for aflatoxins M1 has the advantages of being high in sensitivity, quick and convenient in detection, high in accuracy, good in repeatability and strong in specificity.

Description

technical field [0001] The invention belongs to the technical field of immunological detection and analysis, relates to food safety detection technology, in particular to a kind of aflatoxin M 1 Chemiluminescent detection kit and preparation method thereof. Background technique [0002] Aflatoxins are mainly produced by Aspergillus flavus ( Asperillus Flavus ) and Aspergillus parasitica ( Asperillus Parasiticus ) produced metabolites, mainly B 1 , B 2 , G 1 , G 2 , M 1 and M 2 . In 1993, aflatoxin was classified as a Class I carcinogen by the Cancer Research Institute of the World Health Organization (WHO). Aflatoxin M 1 (Aflatoxin M 1 ) was first discovered by Alleroft in 1963 as aflatoxin B 1 hydroxylated metabolites. Ingestion of aflatoxin B by mammals 1 Aflatoxin B in the body of contaminated feed or food 1 Hydroxylation to aflatoxin M 1 , Aflatoxin M 1 A part will be excreted from urine and milk, and a part will remain in the edible parts of animals, ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N21/76
CPCG01N21/76
Inventor 刘丽青曹晶常燕胡雪婷杜爱铭徐兵
Owner 太原瑞盛生物科技有限公司
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