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Dual-output sensor for lysozyme detection and preparation method of lysozyme

A dual-output, lysozyme technology, applied in the field of analytical chemistry, can solve the problems of complicated operation, limited application, a lot of time, etc., and achieve the effect of simple operation, obvious characteristics and low cost.

Inactive Publication Date: 2013-10-09
FUZHOU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Since lysozyme usually takes a long time to combine with its aptamer DNA, these methods require a lot of time, and the operation is relatively complicated and the cost is high, which greatly limits its application in field detection and daily life.

Method used

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  • Dual-output sensor for lysozyme detection and preparation method of lysozyme
  • Dual-output sensor for lysozyme detection and preparation method of lysozyme
  • Dual-output sensor for lysozyme detection and preparation method of lysozyme

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Embodiment 1

[0021] According to references (Sivakumar, K.; Xie, F.; Cash, B. M.; Long, S.; Barnhill, H. N.; Wang, Q. A Fluorogenic 1,3-Dipolar Cyclodition Reaction of 3-Azidocoumarins and Acetylenes. Org. Lett ., 2004, 6, 4603-4606.) Preparation of 3-azido-7-hydroxycoumarin compound. The specific steps are as follows: Weigh 1.38 g of 2,4-dihydroxybenzaldehyde and 1.12 g of N-acetylglycine into 50 mL of anhydrous sodium acetate, and ultrasonically dissolve, then reflux and stir for 4 hours. After the reaction is completed, the resulting mixture is poured into ice water to obtain a yellow precipitate, filter the yellow precipitate, and wash the precipitate with ice water, and then reflux in 30 mL of a mixture of hydrochloric acid and ethanol (2:1) for 1 hour. After the reaction is complete, use an ice bath to dissolve the above Cool the solution, and dilute the above solution with 20 mL of ice water; add 20 mmol of sodium nitrite, stir for 10 minutes, then add 30 mmol of sodium azide, and s...

Embodiment 2

[0023] The preparation of the gold nanoparticle solution functionalized by triazole coumarin compound, the specific steps are as follows:

[0024] (1) According to the reference (Liu, D.; Chen, W.; Wei, J.; Li, X.; Wang, Z.; Jiang, X. A Highly Sensitive, Dual-Readout Assay Based on Gold Nanoparticles for Organophosphorus and Carbamate Pesticides. Anal. . 2012 , 84 , 4185-4191.) To prepare a gold nanoparticle solution with a diameter of 13 nm, the specific steps are as follows: first, clean the glassware such as the round bottom flask and reflux tube used in the preparation process with aqua regia, and then add chloroauric acid (0.206 mg / mL) into 25 mL water, stirred, heated to reflux, after boiling, added 1.0 mL of 1.3% sodium citrate solution to the above-mentioned hot water solution, continued to heat and reflux, and stirred vigorously for 15 minutes. During this process, the color of the solution changed from light red to Turned into wine red. Finally, stop the solutio...

Embodiment 3

[0028] Preparation method of dual-output sensor for detection of lysozyme: adding different concentrations of lysozyme to the solution of gold nanoparticles functionalized with triazole coumarin compound, shaking on a shaker for 30 minutes, and observing the gold nanoparticles with the naked eye The color of the solution gradually changed from wine red to blue-purple, and the process was accompanied by the recovery of fluorescence, which were detected by fluorescence spectrum and ultraviolet spectrum respectively. It was found that the fluorescence intensity of the solution increased gradually with the increase of the concentration of lysozyme, and the absorbance at 526 nm gradually decreased, and the absorption wavelength gradually shifted to the long-wave direction.

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Abstract

The invention discloses a dual-output sensor for lysozyme detection as well as a preparation method and an application of the lysozyme. The preparation method of the lysozyme comprises the following steps of: (1), preparing 3-azide-7 hydroxycoumarin; (2), mixing 1.0 mM of propargylamine with 1.0 mM of 3-azide-7 hydroxycoumarin, adding 0.1 mM of CuSO4 and 0.5 mM of sodium ascorbate, cultivating for 10 minutes at the room temperature, generating click chemical reaction to generate a triazole coumarin compound; (3), preparing a gold nanoparticle solution with diameter of 13 nm; and (4), mixing 10mu M of the riazole coumarin compound with 100mu L of the gold nanoparticle solution for two minutes in 10 mM of 3-(N-morpholinyl) propanesulfonic acid buffer solution with a pH value of 7.5, and adding 0.25 mM of polyvinylpyrrolidone. According to the visual inspection, the sensor can be used for detecting the lysozyme with the lowest concentration of 50ng / mL and has been successfully applied to detecting the content of the lysozyme in egg plain white. The dual-output sensor disclosed by the invention has the advantages of being simple to operate, low in cost, high in sensitivity and good in specificity.

Description

technical field [0001] The invention belongs to the field of analytical chemistry, and in particular relates to a dual-output sensor for lysozyme detection and a preparation method thereof. Background technique [0002] The full name of lysozyme is 14-O-lysozyme, also known as N-acetylcytate hydrolase, and its isoelectric point pI is about 11. It is an alkaline hydrolase that exists in the tissues and body fluids of microorganisms and various animals and plants. middle. It can cleave the -14-glycoside bond between N-acetylmuramic acid and N-acetylglucosamine in bacterial cells, so it has antibacterial, anti-tumor, and angiogenesis-inhibiting activities, and plays an important role in organisms. Mastitis is one of the three major diseases of dairy cows, which has a great impact on the dairy industry. The low content of lysozyme in natural cow milk may be the main reason for the high incidence of mastitis. At the same time, lysozyme is also closely related to various tumor d...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N21/25G01N21/78G01N21/64G01N21/33
Inventor 林振宇邱素艳郭隆华邱彬陈国南
Owner FUZHOU UNIV
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