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Enzyme response type probe based on rare earth metal and preparation method and application thereof

A kind of rare earth metal, rare earth metal salt technology

Pending Publication Date: 2021-09-07
FUJIAN INST OF RES ON THE STRUCTURE OF MATTER CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, organic dyes, quantum dots, and metal-organic framework materials used in traditional fluorescence analysis methods have problems such as poor photochemical stability, high toxicity, high cost, and long time consumption, and cannot avoid the interference of complex biomolecules in the sample to be tested, making these The method cannot realize the rapid and accurate direct detection of alkaline phosphatase concentration

Method used

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  • Enzyme response type probe based on rare earth metal and preparation method and application thereof
  • Enzyme response type probe based on rare earth metal and preparation method and application thereof
  • Enzyme response type probe based on rare earth metal and preparation method and application thereof

Examples

Experimental program
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Effect test

preparation example 1

[0046] Preparation Example 1: Synthesis of Rare Earth Metal-Based Enzyme-Responsive Probes (Dual Ion Probes)

[0047] The probe is prepared by stirring at room temperature, using rare earth nitrates and ATP compounds as raw materials, and the preparation method is as follows:

[0048] (1) Weigh the same amount of Ce(NO 3 ) 3 ·6H 2 O, Tb(NO 3 ) 3 ·6H 2 O is dissolved in the Tris-HCl aqueous solution of pH=9.0, is prepared into the rare earth ion solution that the total concentration of rare earth ion is 4mM;

[0049] (2) ATP was weighed and dissolved in Tris-HCl aqueous solution to prepare an ATP solution with a concentration of 2mM;

[0050] (3) Take 1 mL of the rare earth ion solution in step (1) and mix evenly, add dropwise to 1 mL of the ATP solution in step (2), and stir at room temperature for one minute.

[0051] The characterization results of the prepared probes (also known as dual ion probes) are shown in figure 2 , wherein, (a), (b) are transmission electron...

Embodiment 1

[0064] Embodiment 1: the detection of alkaline phosphatase concentration

[0065] (1) Using a polystyrene 96-well plate as the carrier used for detection, add 100 μL of the probe solution prepared in Preparation Example 1 in 7 rows in sequence to the set microwells, and the concentrations in each row are 32, 16, 8, 4, 2, 1, 0.5mM, add 100μL alkaline phosphatase (ALP) aqueous solution with different concentrations in each column in turn, the concentrations are 3000, 2500, 2000, 1500, 1250, 1000, 750, 500, 250, 125, 62.50, 31.25, 15.63, 7.81, 0U / L; place at 37°C for 3 hours and shake at a constant temperature, measure the fluorescence intensity of the mixed solutions in 7 rows and 15 columns, and calculate the fluorescence quenching efficiency of each group of mixed solutions; when the fluorescence quenching efficiency is the largest The probe concentration corresponding to the mixed solution is 4mM, and the concentration of alkaline phosphatase in the mixed solution is used for...

Embodiment 2

[0068] Example 2: The detection specificity verification of alkaline phosphatase to be tested

[0069] (1) The reagents, instruments, and probes required for the experiment are all consistent with those in Example 1.

[0070] (2) Common interfering substances in the blood were selected for the experiment: arginine, glycine, biotin, citric acid, lactose, bovine serum albumin (BSA), glucose oxidase (GOD), cholinesterase, metal ions (K + , Ca 2+ , Cl - 、Na + , Mg 2+ ).

[0071] (3) After grouping the detection well plate in the 96-well plate, add 100 μL of the above-mentioned interfering substance aqueous solution with a concentration of 100 mg / mL and 100 μL of an alkaline phosphatase aqueous solution with a concentration of 100 U / L to 100 μL containing the probe ( In the Tris-HCl buffer solution with a final concentration of 4 mM) at pH = 9.0, the 96-well plate was placed at 37°C for a constant temperature shaking reaction for 3 hours, and the reacted 96-well plate was plac...

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Abstract

The invention discloses an enzyme response type probe based on rare earth fluorescence resonance energy transfer and a preparation method and application thereof in alkaline phosphatase detection. The enzyme response type probe utilizes supramolecular self-aggregation of rare earth sensitized ions, rare earth activated ions and phospholipid molecules, effectively shortens the energy transfer distance between the rare earth sensitized ions and the activated ions, successfully constructs a nano probe with good fluorescence characteristics, and realizes specific and high-sensitivity detection of alkaline phosphatase. The enzyme response type probe provided by the invention is simple and quick to prepare, safe and non-toxic, and can effectively avoid interference in a biological complex system. The detection of the concentration of alkaline phosphatase in serum or whole blood can be realized only by simply mixing the nano probe with a sample to be detected in the detection process, the advantages of simplicity, sensitivity, good specificity, economy, practicability and the like are achieved, and a simple and efficient new method and corresponding technical support are provided for the detection of alkaline phosphatase in a complex system.

Description

technical field [0001] The invention relates to the technical field of biological detection, in particular to a rare-earth metal-based enzyme-responsive probe and its preparation method and application. Background technique [0002] Enzymes are catalysts for the metabolism of organisms and play an important role in maintaining the normal operation of various biochemical and physiological reactions of organisms. With the further understanding of enzymes, it has been found that biological enzymes are closely related to cell activities, pathological reactions, and disease occurrence, and most diseases are related to enzyme deficiency or synthesis disorders. Therefore, enzymes play an indicative role in disease prevention, early diagnosis, and drug response. Alkaline phosphatase is an important enzyme disease marker, which is closely related to the occurrence and development of major diseases such as cancer, bone disease, liver and gallbladder disease, and diabetes. The concen...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07H23/00C09K11/06G01N21/64B82Y30/00B82Y40/00
CPCC07H23/00C09K11/06G01N21/6428B82Y30/00B82Y40/00G01N2021/6432G01N2021/6417C09K2211/182
Inventor 刘䶮陈学元杨颖婕张云钦李幸俊高航
Owner FUJIAN INST OF RES ON THE STRUCTURE OF MATTER CHINESE ACAD OF SCI
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