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Acetylcholin esterase (AChE) catalytic spectrum sensing detection system and detection method

A detection method and sensing technology, applied in the detection field, can solve the problems of oxidation potential interference, background interference, low accuracy, etc.

Pending Publication Date: 2021-10-22
HUAQIAO UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] Among the above AChE activity detection technologies, the Ellman detection method has the disadvantages of background interference, cumbersome process and high false positive signal; the fluorescence "turn on" method is not only time-consuming and costly, but also the fluorescence method is difficult to be suitable for real-time and rapid on-site monitoring; Electrochemical detection instruments are easy to operate, fast in measurement speed, and easy to carry, and have attracted widespread attention. At present, they mainly focus on potentiometric and amperometric AChE biosensors, but the oxidation potential is too high and the interference is serious, resulting in low accuracy and poor sensitivity.

Method used

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  • Acetylcholin esterase (AChE) catalytic spectrum sensing detection system and detection method
  • Acetylcholin esterase (AChE) catalytic spectrum sensing detection system and detection method
  • Acetylcholin esterase (AChE) catalytic spectrum sensing detection system and detection method

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] Embodiment 1: Au NPs-(S-C 16 -TACN)-Zn 2+ Preparation of Nano-Au Catalysts

[0042] Preparation of Au NPs-(S-C 16 -TACN)-Zn 2+ The method for the nanometer Au type catalyst specifically comprises the following steps:

[0043] 1) Synthesis of Au NPs with a particle size of about 3nm: Referring to the method of reducing chloroauric acid, the TEM of the prepared 3nm Au NPs is as follows: figure 2 shown.

[0044] 2) Synthesis of sulfhydryl molecule HS-C with a TACN ring at the end 16 -TACN steps are as follows:

[0045] ①The synthesis steps of Boc-protected TACN are as follows:

[0046] Weigh 0.3204g 1,4,7-triazacyclononane (TACN) (2.48mmol) with an analytical balance, and dissolve it in a flask filled with 20mL of chloroform, add 230μL tri Ethylamine. Continue to stir and remove the air in the flask, weigh 1.082g di-tert-butyl dicarbonate (4.96mmol) with an analytical balance, and dissolve it in a flask containing 20mL of chloroform, and slowly add it dropwise to...

Embodiment 2

[0055] Embodiment 2: Au NPs-(S-C 16 -TACN)-Zn 2+ Hydrolysis experiment of substrate molecule HPNP catalyzed by the nano-Au catalyst

[0056] Au NPs-(S-C 16 -TACN)-Zn 2+ The experimental steps of the nano-Au type catalyst catalyzing the hydrolysis reaction of the substrate molecule HPNP are as follows:

[0057] Add 160 μL of 1 mM HPNP solution to 1840 μL of Au NPs-(S-C 16 -TACN)-Zn 2+ In the 1mM HEPES buffer (pH 7.24) of the nano-Au catalyst, the HEPES buffer solution was adjusted to the required pH by NaOH solution, shaken, the mixture was heated at 30°C for 0.5h, and its UV-vis absorption spectrum was scanned immediately .

Embodiment 3

[0058] Embodiment 3: utilize AuNPs-(S-C 16 -TACN)-Zn 2+ The detection of AChE using nano-Au-type catalysts

[0059] A novel AChE catalytic spectral sensing detection method, comprising the following steps:

[0060] (1) Sample preparation:

[0061] 1) With reference to the method of Example 1, AuNPs-(S-C 16 -TACN)-Zn 2+ Nano-Au catalysts.

[0062] 2) AChE solution preparation: first prepare HEPES buffer solution, dissolve AChE (commercially available product) after filtering through a filter membrane and store it at low temperature, and dilute it with bovine serum albumin solution to the required activity when used.

[0063] (2) Detection: AChE with different activities was added to Au NPs-(S-C 16 -TACN)-Zn 2+ In the mixed solution system of nanometer Au type catalyst and HPNP, react at a certain temperature for several minutes, scan its UV-vis absorption spectrum immediately, and calculate the content of AChE in the sample solution through the change of absorbance at 29...

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Abstract

The invention discloses an acetylcholin esterase (AChE) catalytic spectrum sensing detection system and a detection method. A nano Au-type AChE catalytic spectrum sensing system is constructed by using HPNP as a substrate of a catalytic reaction by utilizing the characteristic that a substrate molecule 2-hydroxypropyl-4-nitrophenyl phosphodiesters (HPNP) is similar to an organophosphorus pesticide structure and utilizing the characteristic that AChE and an organophosphorus compound are subjected to a phosphorylation reaction, the detection limit of AChE activity is 0.012 U.L<-1> (3 sigma / k, n is equal to 11, and k is the slope of a relation curve), and the detection limit of AChE activity is 0.012 U.L<-1> (3[sigma] / k, n is equal to 11), and the linear range is 0-6U. L<-1>.

Description

technical field [0001] The invention belongs to the technical field of detection, and in particular relates to the establishment and detection technology of a novel acetylcholinesterase catalytic spectrum sensing detection system. Background technique [0002] Acetylcholinesterase (AChE) is an important natural enzyme in the process of biological nerve transmission. It is ubiquitous in the peripheral nervous system and can specifically catalyze the hydrolysis reaction of the substrate acetylcholine to quickly hydrolyze it into choline and acetate. . This enzymatic reaction process is associated with the occurrence of a series of diseases, such as Alzheimer's disease (AD), inflammation and poisoning by nerve agents. At present, studies have shown that the introduction of AChE inhibitors can increase the level of neurotransmitter acetylcholine, so as to alleviate the onset of AD and improve AD symptoms. Therefore, the development of analytical and detection methods for AChE ...

Claims

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Application Information

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IPC IPC(8): G01N21/31G01N21/33G01N31/10
CPCG01N21/31G01N21/33G01N31/10
Inventor 沈江珊姚美霞曹荧娟
Owner HUAQIAO UNIVERSITY
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