Supermolecular assembly for trypsin based on label-free fluorescence detection and preparation method of supermolecular assembly

A supramolecular assembly and fluorescence detection technology, applied in the interdisciplinary fields of materials, biology and analytical chemistry, can solve problems such as unseen gemini surfactants, and achieve the effects of good salt resistance, extended application and low cost

Active Publication Date: 2017-10-10
TAIYUAN UNIV OF TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Through the search, there are no patents and reports on the use of gemini surfactants for fluorescence detection

Method used

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  • Supermolecular assembly for trypsin based on label-free fluorescence detection and preparation method of supermolecular assembly
  • Supermolecular assembly for trypsin based on label-free fluorescence detection and preparation method of supermolecular assembly
  • Supermolecular assembly for trypsin based on label-free fluorescence detection and preparation method of supermolecular assembly

Examples

Experimental program
Comparison scheme
Effect test

Embodiment approach 1

[0024] (1) Preparation of supramolecular assembly solution

[0025] Ⅰ Determination of critical micelle concentration CMC of quaternary ammonium cationic gemini surfactant;

[0026] The quaternary ammonium cationic surfactant used is ethylene didodecyl dimethyl ammonium bromide, and the structural formula is as follows:

[0027]

[0028] Dissolve ethylenedidodecyldimethylammonium bromide in a phosphate buffer solution with a concentration of 10mM and a pH of 8.0 to prepare 2mL with a concentration range of 10 -2 ~10 -7 mol L -1 Gradient solution of Nile Red; use a microsampler to pipette 10 μL of Nile Red in acetone solution and add it to the gradient solution so that the final concentration of Nile Red in the gradient solution is 10 -6 mol L -1 , with 100kHz exposure ultrasonic treatment for 10 minutes, after standing still for 0.5 hours, carry out fluorescence emission spectrum detection; take the emission peak intensity at 635nm as the ordinate, and the logarithmic c...

Embodiment approach 2

[0044] (1) Preparation of supramolecular assembly solution

[0045] Ⅰ Determination of critical micelle concentration CMC of quaternary ammonium cationic gemini surfactant;

[0046] The quaternary ammonium cationic surfactant used is ethylene ditetradecyl dimethyl ammonium bromide, and the structural formula is as follows:

[0047]

[0048] Dissolve ethylene ditetradecyl dimethyl ammonium bromide in a phosphate buffer solution with a concentration of 10 mM and pH 8.0, and prepare 2 mL of a concentration range of 10 -2 ~10 -7 mol L -1 Gradient solution of Nile Red; use a microsampler to pipette 30 μL of Nile Red in acetone solution and add it to the gradient solution so that the final concentration of Nile Red in the gradient solution is 10 -6 mol L -1 , with a 70kHz exposure ultrasonic treatment for 30 minutes, and after standing for 2 hours, perform fluorescence emission spectrum detection; take the emission peak intensity at 635nm as the ordinate, and the logarithmic ...

Embodiment approach 3

[0061] (1) Preparation of supramolecular assembly solution

[0062] Ⅰ Determination of critical micelle concentration CMC of quaternary ammonium cationic gemini surfactant;

[0063] The quaternary ammonium cationic surfactant used is ethylene dihexadecyl dimethyl ammonium bromide, and the structural formula is as follows:

[0064]

[0065] Dissolve ethylenedihexadecyldimethylammonium bromide in a phosphate buffer solution with a concentration of 10mM and a pH of 8.0, and prepare 2mL with a concentration range of 10 -2 ~10 -7 mol L -1 Gradient solution of Nile Red; use a microsampler to pipette 50 μL of Nile Red in acetone solution and add it to the gradient solution so that the final concentration of Nile Red in the gradient solution is 10 -6 mol L -1 , with a 40kHz exposure ultrasonic treatment for 60 minutes, and after standing for 12 hours, the fluorescence emission spectrum detection is carried out; the emission peak intensity at 635nm is taken as the ordinate, and ...

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Abstract

The invention relates to a supermolecular assembly for trypsin based on label-free fluorescence detection and a preparation method of the supermolecular assembly and belongs to the technical field of interdisciplinary science of materials, biology and analytical chemistry. The supermolecular assembly is characterized by being compound aqueous solution formed by a quaternary ammonium cationic gemini surfactant and heparin sodium under the electrostatic action. The preparation method comprises the following steps: firstly determining critical micelle concentration of the gemini surfactant, forming the supermolecular assembly by the gemini surfactant and the heparin sodium at the concentration, and enabling a hydrophobic dye Nile red to enter the inner cavity of the assembly, so that fluorescence emission is enhanced; then adding protamine which has specific interaction with the heparin sodium, and disassembling, so that the fluorescence emission intensity is reduced with increase of the concentration of the protamine; and then adding trypsin to hydrolyze the protamine and restore the assembly, so that the fluorescence emission intensity is increased with increase of trypsin concentration, and trypsin activity detection is realized. The supermolecular assembly provided by the invention is simple to prepare and low in cost and has good salt resistance and a broad application prospect.

Description

technical field [0001] The invention discloses a supramolecular assembly for unlabeled fluorescent detection of trypsin and a preparation method thereof, belonging to the technical field of interdisciplinary materials, biology and analytical chemistry. Background technique [0002] Trypsin is one of the most important digestive enzymes in protein decomposition. The secretion, activation, inhibition and circulation imbalance of trypsinogen can lead to acute or chronic pancreatic diseases, such as pancreatic cancer. Approximately 35,000 cases of pancreatic cancer each year are caused by an imbalance of serine proteases, especially trypsin. In addition, trypsin not only acts as a digestive enzyme, but also restricts the decomposition of precursors of other enzymes such as chymotrypsinogen, carboxypeptidase, and phospholipase, and activates them. [0003] Currently developed methods for the determination of trypsin mainly include UV spectrophotometry, colorimetry, fluorescence,...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N21/64
Inventor 贾兰陈松高菲张耀东侯文娟
Owner TAIYUAN UNIV OF TECH
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