Alkaline phosphatase determination method based on carbon-dots fluorescence ''quenching-recovery''

A technology of carbon dot fluorescence and determination method, which is applied in the field of biological analysis and detection of nanomaterials, and can solve problems such as analysis and determination that have not yet been found.

Active Publication Date: 2016-12-21
JIANGNAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

As far as we know, there is no report on the use of redox to reg

Method used

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  • Alkaline phosphatase determination method based on carbon-dots fluorescence ''quenching-recovery''
  • Alkaline phosphatase determination method based on carbon-dots fluorescence ''quenching-recovery''
  • Alkaline phosphatase determination method based on carbon-dots fluorescence ''quenching-recovery''

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Experimental program
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Embodiment 1

[0017] a. Weigh 1.1g of ascorbic acid and dissolve it in 25mL of ethanol solution, mix it with 25mL of ultrapure water, stir to form a uniform solution, transfer it to the reaction kettle, heat at 180°C for 4h, and cool the obtained dark brown solution with dichloro Methane was extracted, and the obtained solution was dialyzed with ultrapure water for 24 hours as a fluorescent probe;

[0018] b. Add 50 μL 0.1mol / L trisodium ascorbic acid solution and 50 μL alkaline phosphatase solution of different concentrations into 400 μL Tris-HCl buffer solution with pH=8.5, and add 100 μL potassium permanganate (0.01 mmol / L) and 0.35mg / mL carbon dot mixture (the fluorescence intensity is recorded as F 0 ), and measure the fluorescence intensity after 2 min of reaction.

Embodiment 2

[0020] a. Add 25mL of milk to 20mL of ultrapure water, stir it into a uniform solution, transfer it to a reaction kettle, heat it at 180°C for 2h, filter it with a microporous filter after cooling, and dialyze the obtained solution with ultrapure water for 24h as Fluorescent probes;

[0021] b. Add 50 μL 0.1mol / L p-aminophenylphosphate sodium salt solution and 50 μL alkaline phosphatase solution of different concentrations to 400 μL Tris-HCl buffer solution with pH=8.5, react for 25 minutes and add 100 μL containing potassium permanganate ( 0.01mmol / L) and 0.35mg / mL carbon dot mixture (the fluorescence intensity is recorded as F 0 ), and measure the fluorescence intensity after 2 min of reaction.

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Abstract

The invention provides a sensitive detection method of alkaline phosphatase based on carbon-dots fluorescence ''quenching-recovery''. Carbon dots prepared by a simple hydrothermal method have low toxicity and good water solubility and fluorescent property. In the presence of potassium permanganate, fluorescence of carbon dots is quenched; and with a hydrolysate ascorbic acid or p-aminophenol of alkaline phosphatase (ALP), quenched fluorescence will be recovered again. By high catalytic activity of ALP, a novel fluorescence analysis method is established to realize highly sensitive detection of ALP. The method has advantages of novel principle, high sensitivity, good selectivity and simple and fast operation, and is adopted to avoid interference of false signals in a fluorescence quenching method and widen the application range of carbon dots-based biological detection.

Description

Technical field: [0001] The invention relates to the field of biological analysis and detection of nanomaterials, in particular to the application of novel fluorescent carbon nanomaterials-carbon dots in the detection of alkaline phosphatase activity. Background technique: [0002] Alkaline phosphatase (ALP) is a membrane-bound enzyme that widely exists in biological tissues, and it is abundant in liver, bone and kidney. It has a wide range of substrate specificity and can catalyze various phosphate compounds. hydrolysis. Abnormal levels of ALP in serum are closely related to various diseases such as skeletal diseases, diabetes and hepatic insufficiency [Colombatto P.; Randone A.; Civitico G.; Gorin J.M.; Dolci L.; Medaina N.; Oliveri F.; Verme G.; Marchiaro G.; Pagni R.; Karayiannis P.; Thomas H.C.; Hess G.; Bonino F.; In addition, ALP is also one of the commonly used labeling reagents in biological research. When used as a marker for enzyme-linked immunoassay, it has the...

Claims

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Application Information

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IPC IPC(8): G01N21/64
CPCG01N21/6486
Inventor 王光丽方馨吴秀明李在均
Owner JIANGNAN UNIV
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