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Method for determining aflatoxin B1 based on immunomagnetic beads

A technology of aflatoxin and immunomagnetic beads, which is applied in the direction of measuring devices, instruments, scientific instruments, etc., and can solve the problems that the quantitative determination of mycotoxins by immunomagnetic beads and flow cytometry has not been reported.

Pending Publication Date: 2020-06-12
HAINAN UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] At present, the application of immunomagnetic separation technology and flow cytometry is more common in the field of medical analysis and testing, but there is no report on the application of immunomagnetic beads and flow cytometry to the quantitative determination of mycotoxins

Method used

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  • Method for determining aflatoxin B1 based on immunomagnetic beads

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Experimental program
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Effect test

Embodiment 1

[0062] Precisely pipette 1mg of 0.5μm carboxyl magnetic beads into a 1.5mL centrifuge tube, then wash 3 times with 0.1M MES buffer (pH6.2), then perform magnetic separation, and resuspend the carboxyl magnetic beads after magnetic separation in PBS buffer to prepare magnetic bead suspension. Add 0.77mg EDC and 1.09mgsulfo-NHS to the carboxyl magnetic bead suspension and incubate for 20 minutes at 25°C on a Limber BE-1100 mixer to activate the carboxyl groups on the surface of the magnetic beads, and centrifuge to obtain activated carboxyl groups magnetic beads. Then, resuspend the above-mentioned activated carboxyl magnetic beads in 0.5ml PBS buffer (containing 100μl AFB 1 -BSA) and shake gently on its Limbell BE-1100 mixer for 1 h to form AFB at 25 °C 1 -BSA-MBs solution. After magnetic separation, the AFB 1 -BSA-MBs were stored in 10ml PBS buffer containing 0.05% Tween-20 and 1% BSA, AFB 1 -BSA-MBs preservation solution was diluted to 1 / 100 with PBS buffer to obtain AFB...

Embodiment 2

[0064]Precisely pipette 1mg of 0.5μm carboxyl magnetic beads into a 1.5mL centrifuge tube, then wash 3 times with 0.1M MES buffer (pH6.2), then perform magnetic separation, and resuspend the carboxyl magnetic beads after magnetic separation in PBS buffer to prepare magnetic bead suspension. Add 0.77mg EDC and 1.09mgsulfo-NHS to the carboxyl magnetic bead suspension and incubate for 20 minutes at room temperature (25°C) on its Limber BE-1100 mixer to activate the carboxyl groups on the surface of the magnetic beads, centrifuge, Get activated carboxyl magnetic beads. Then, resuspend the above-mentioned activated carboxyl magnetic beads in 0.5ml PBS buffer (pH7.4, containing 100μL AFB 1 -BSA) and shake gently for 1 h on its Limbell BE-1100 mixer to form AFB at room temperature (25°C). 1 -BSA-MBs solution. After magnetic separation, the AFB 1 -BSA-MBs were stored in 10ml PBS buffer containing 0.05% Tween-20 and 1% BSA, AFB 1 -BSA-MBs preservation solution was diluted to 1 / 100...

Embodiment 3

[0066] Precisely pipette 1mg of 0.5μm carboxyl magnetic beads into a 1.5mL centrifuge tube, then wash 3 times with 0.1MMES buffer (pH6.2), then perform magnetic separation, and resuspend the carboxyl magnetic beads after magnetic separation in PBS buffer In , the magnetic bead suspension was prepared. Add 0.77 mg EDC and 1.09 mg sulfo-NHS to the carboxyl magnetic bead suspension and incubate for 20 minutes at room temperature (25°C) on its Limber BE-1100 mixer to activate the carboxyl groups on the surface of the magnetic beads, centrifuge, Get activated carboxyl magnetic beads. Then, resuspend the above-mentioned activated carboxyl magnetic beads in 0.5ml PBS buffer (pH7.4, containing 100μL AFB 1 -BSA) and shake gently for 1 h on its Limbell BE-1100 mixer to form AFB at room temperature (25°C). 1 -BSA-MBs solution. After magnetic separation, the AFB 1 -BSA-MBs were stored in 10ml PBS buffer containing 0.05% Tween-20 and 1% BSA, AFB 1 -BSA-MBs preservation solution was di...

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Abstract

The invention belongs to the technical field of aflatoxin determination methods, and provides a method for determining aflatoxin B1 based on immunomagnetic beads. The determination method comprises the steps of activating the magnetic beads, modifying the magnetic beads with antigens, preparing an immunomagnetic bead system, carrying out flow cytometry detection and the like. According to the method for determining aflatoxin B1 based on immunomagnetic beads, the pretreatment method is conventional and concise, is clear in operation, high in instrument automation degree and sensitive and accurate in analysis, a large number of detection results can be rapidly obtained in real time on the basis of a flow cytometer, the detection limit of the concentration of aflatoxin B1 is low, and the detection limit can reach 0.2034-0.3107 microgram / L.

Description

technical field [0001] The present invention relates to the technical field of aflatoxin assay method, in particular to a kind of aflatoxin B based on immunomagnetic beads 1 method of measurement. Background technique [0002] Aflatoxins (AFT) are a class of toxins produced by Aspergillus flavus and Aspergillus parasiticus with various derivatives. Among them, aflatoxin B 1 (AFB 1 ) is the most toxic and carcinogenic. Recent studies have found that AFB 1 Is a causative agent of hepatocellular carcinoma, growth inhibition, modulation of the immune system, and malnutrition. It is highly toxic to humans and animals, equivalent to 10 times that of potassium cyanide. Furthermore, it can also be transferred between organisms through the food chain, thereby posing a great threat to the metabolic cycle. Pregnant and lactating women who have aflatoxin metabolites or biomarkers in breast milk, serum, or cord blood will reduce human fertility. [0003] AFB 1 and its producing ...

Claims

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Application Information

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IPC IPC(8): G01N33/577G01N33/531
CPCG01N33/577G01N33/531
Inventor 陈健宿瑞奇姚广龙曹献英唐雪梅冯露
Owner HAINAN UNIVERSITY
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