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Method for detecting acetyl choline by fluorescence ratio

A technology of acetylcholine and acetylcholinesterase, applied in fluorescence/phosphorescence, measuring device, material analysis by optical means, etc., can solve the problems of complicated operation method, long reaction time, low sensitivity, etc., and achieve high fluorescence luminescence intensity and sensitivity High, fast and sensitive detection effect

Inactive Publication Date: 2019-07-23
南宁师范大学
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, most methods have defects such as complicated operation methods, long reaction time, and low sensitivity.

Method used

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  • Method for detecting acetyl choline by fluorescence ratio
  • Method for detecting acetyl choline by fluorescence ratio
  • Method for detecting acetyl choline by fluorescence ratio

Examples

Experimental program
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Effect test

Embodiment 1

[0024] A method for fluorescence ratio detection of acetylcholine, comprising the following steps:

[0025] Step 1. Prepare equal volumes of multiple standard solutions containing the same concentration of acetylcholinesterase, choline oxidase, horseradish peroxidase, o-phenylenediamine and carbon quantum dots, and containing different concentrations of acetylcholine, specifically: A , with acetylcholine dry powder and ultrapure water to prepare a concentration of 1 × 10 -2 The stoste of mol / L; B, the Tris-HCl damping solution that B, described stoste is 7.4 with pH, ​​concentration is 0.05mol / L carries out gradient dilution, makes the acetylcholine standard solution of different concentrations, and its concentration is successively 0mol / L, 5×10 -5 mol / L, 1×10 -4 mol / L, 1.5×10 -4 mol / L, 2×10 -4 mol / L; C. Add 100 μl of acetylcholinesterase solution with a concentration of 80 U / L and 100 μl of choline oxidase solution with a concentration of 4 U / mL to 100 μl of the acetylcho...

Embodiment 2

[0029] A method for fluorescence ratio detection of acetylcholine, the process is roughly the same as in Example 1, the difference is: in C, specifically: first add the same volume of acetylcholinesterase solution, choline oxidation Enzyme solution, horseradish peroxidase solution and o-phenylenediamine solution were mixed at 37°C for 30 minutes, then the same volume of carbon quantum dot solution was added respectively, stirred with a micro-injector, and mixed and reacted at room temperature for 1 minute. That is obtained; the fluorescence spectrograms of the multiple standard solutions are as follows: figure 1As shown, among them, I 442 is the fluorescence of carbon dots, I 573 is the fluorescence of 2,3-diaminophenazine, and the concentration of acetylcholine standard solution corresponding to curve a, b, c, d, e is 0mol / L, 5×10 -5 mol / L, 1×10 - 4 mol / L, 1.5×10 -4 mol / L, 2×10 -4 mol / L, take the concentration of acetylcholine as the abscissa, I 573 / I 442 As the ordi...

Embodiment 3

[0032] A method for fluorescence ratio detection of acetylcholine, the process of which is roughly the same as that of Example 2, the difference being that when the carbon quantum dots are prepared, before the mixed solution is transferred to the reaction kettle, it also includes putting the mixed solution into In a container with a micro-current generator, conduct electrification treatment with a current of 10mA and a treatment time of 4min.

[0033] Adopt the method of the present embodiment to be 6.35 * 10 to the detection limit of acetylcholine -6 mol / L.

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PUM

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Abstract

The invention discloses a method for detecting acetyl choline by a fluorescence ratio. The method comprises the following steps of step1, preparing multiple standard solutions with the same volume, having the same concentrations of acetylcholin esterase, choline oxidase, horse radish peroxidase, o-phenylenediamine and carbon quantum dots, and having the different concentrations of acetyl choline;step 2, detecting the fluorescence intensities I442 and I573 of the multiple standard solutions at 442nm and 573nm, and building a linear relation between the I573 / I442 and the acetyl choline concentration; and step 3, preparing a solution to be detected with the unknown acetyl choline concentration, detecting the fluorescence intensities I442 and I573 of the solution at 442nm and 573nm, and thuscomputing the acetyl choline concentration in the solution to be detected according to the linear relation acquired in the step 2. The method provided by the invention has the advantages of simple operation, quick detection, high sensitivity and the like.

Description

technical field [0001] The invention relates to the technical field of acetylcholine detection. More specifically, the present invention relates to a method for the fluorescence ratiometric detection of acetylcholine. Background technique [0002] Acetylcholine (ACh) is found in neuromuscular junctions and synapses in animals, and abnormally low concentrations of ACh can cause various neuropsychological and neuropsychiatric disorders. In addition to their unique optical properties, carbon dots also show low cytotoxicity and good biocompatibility because they mainly contain carbon elements, which makes carbon dots become the traditional heavy metal quantum dots in biological fields such as biological cell markers, disease Alternatives in diagnostics, bioimaging, biodetection, drug delivery applications. At present, domestic and foreign detection methods for acetylcholine mainly include chemiluminescence, fluorescence, spectrophotometry and electrochemical methods. However,...

Claims

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Application Information

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IPC IPC(8): G01N21/64
CPCG01N21/6428G01N2021/6439
Inventor 黄珊肖琦姚建东刘义
Owner 南宁师范大学
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