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Chemiluminescence detection method of ota based on nano-gold labeled aptamer sensor

A technology of chemiluminescence detection and aptasensor, which is applied in the field of analytical chemistry and can solve the problems of poor sensitivity and unstable results of OTA detection methods

Active Publication Date: 2019-05-24
绍兴叁讯科技股份有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The present invention aims at the technical defects of the prior art, and provides an OTA chemiluminescence detection method based on nano-gold labeled aptamer sensors, so as to solve the technical defects of poor sensitivity and unstable results of the OTA detection method in the prior art

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  • Chemiluminescence detection method of ota based on nano-gold labeled aptamer sensor
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  • Chemiluminescence detection method of ota based on nano-gold labeled aptamer sensor

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Embodiment 1

[0041] 1 Experimental principle

[0042] The invention constructs the OTA aptamer sensor on the surface of the magnetic microsphere, uses the carboxyl magnetic microsphere as an immobilized carrier, and utilizes the magnetic separation technology to realize the separation of the aptamer sensor from other heterogeneous phases. In addition, the technical scheme also uses the complementary hybridization between the OTA aptamer and the reporter sequence by modifying the reporter sequence on the surface of the nano-gold particle to label the nano-gold particle as a marker on the surface of the magnetic microsphere. In addition, according to the AuCl formed after the gold nanoparticles are oxidized 4 - The chemiluminescence detection principle of chemiluminescence in the catalytic luminol system, and in this sensing mode, the negative correlation between the OTA concentration and the number of gold nanoparticles labeled on the surface of the magnetic microspheres, thus obtaining th...

Embodiment 2

[0067] An OTA chemiluminescence detection method based on nano-gold labeled aptasensor, comprising the following steps:

[0068] 1) Take the gold nanometer solution and mix it with the thiolated reporter sequence and incubate together, separate the solid and liquid and discard the supernatant to obtain the DNA-AuNPs complex;

[0069] 2) Immobilizing the amino-modified OTA aptamer on the surface of carboxyl magnetic microspheres by aminocarboxyl reaction to obtain a magnetic microsphere-OTA aptamer complex;

[0070] 3) Mix the excess magnetic microsphere-OTA aptamer complex with the sample to be tested and fully react;

[0071] 4) Mix the product obtained in step 3) with excess DNA-AuNPs complex, and take the solid phase after the reaction to obtain a nano-gold-labeled aptamer sensor;

[0072] 5) Mix the gold nanometer-labeled aptasensor described in step 4) with the luminol reagent, and then detect the CL value of the luminescent signal.

[0073] On the basis of the above te...

Embodiment 3

[0081] An OTA chemiluminescence detection method based on nano-gold labeled aptasensor, comprising the following steps:

[0082] 1) Take the gold nanometer solution and mix it with the thiolated reporter sequence and incubate together, separate the solid and liquid and discard the supernatant to obtain the DNA-AuNPs complex;

[0083] 2) Immobilizing the amino-modified OTA aptamer on the surface of carboxyl magnetic microspheres by aminocarboxyl reaction to obtain a magnetic microsphere-OTA aptamer complex;

[0084] 3) Mix the excess magnetic microsphere-OTA aptamer complex with the sample to be tested and fully react;

[0085] 4) Mix the product obtained in step 3) with excess DNA-AuNPs complex, and take the solid phase after the reaction to obtain a nano-gold-labeled aptamer sensor;

[0086] 5) Mix the gold nanometer-labeled aptasensor described in step 4) with the luminol reagent, and then detect the CL value of the luminescent signal.

[0087] On the basis of the above te...

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Abstract

The invention provides an OTA chemiluminiscence detecting method based on a nano-gold label aptasensor. According to the technical scheme adopted by the invention, the OTA chemiluminiscence detecting method comprises the following steps of: taking an OTA aptamer as a stationary phase which is fixedly arranged on a magnetic microsphere surface, taking nano-gold as a label, and modifying a report sequence on the surface of a nano-gold particle, thereby labelling the nano-gold on the aptasensor; based on high affinity of a nucleic acid aptamer, utilizing a competitive relation that OTA competes with the report sequence to combine with the OTA aptamer, enabling the OTA to combine with high-ability report sequence of the aptamer, thereby obtaining a negative correlation between to-be-detected OTA concentration and report sequence amount connected on the sensor; and taking nano-gold connected with the report sequence as a detecting probe, utilizing the principle that a nano-gold catalysis luminal system generates a chemiluminiscence signal to establish corresponding relation between dose of nano-gold connected on the aptasensor with chemical luminous intensity, thereby obtaining quantitative relation between OTA concentration and chemical luminous intensity, and indirectly realizing quantitative analyzing and detecting of OTA. Moreover, detecting sensitivity is greatly improved.

Description

technical field [0001] The present invention relates to the technical field of analytical chemistry, and further relates to nucleic acid aptamer technology, in particular to an OTA chemiluminescence detection method based on a nano-gold labeled aptamer sensor. Background technique [0002] Ochratoxin A (OTA) is a highly toxic mycotoxin, and its toxicity ranks second in the mycotoxin family, second only to aflatoxin. OTA is widely distributed in nature, which not only pollutes grain food, but also seriously pollutes animal food, posing a great threat to human beings. In view of the fact that food safety is related to the safety of the whole society and is a major concern of the country and the people, the establishment of OTA's highly sensitive, simple and rapid analysis method has attracted more and more attention from researchers. [0003] At present, many grass-roots food testing departments mostly use thin-layer chromatography for OTA quantitative detection. This is also...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N21/76
CPCG01N21/76
Inventor 严喜鸾肖义陂刘杰
Owner 绍兴叁讯科技股份有限公司
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