Construction and Application of Novel Fluorescent Polarization Sensor Based on Small Molecule Fluorescent Probe

A reaction and alkyl technology, applied in the field of protein biodetection, to achieve good stability, good water solubility, and high background signal-to-noise ratio

Active Publication Date: 2019-10-29
SHANGHAI NORMAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

These factors make the detection based on the change of fluorescence intensity insufficient in quantitative detection.

Method used

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  • Construction and Application of Novel Fluorescent Polarization Sensor Based on Small Molecule Fluorescent Probe
  • Construction and Application of Novel Fluorescent Polarization Sensor Based on Small Molecule Fluorescent Probe
  • Construction and Application of Novel Fluorescent Polarization Sensor Based on Small Molecule Fluorescent Probe

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Experimental program
Comparison scheme
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specific Embodiment

[0060]

[0061] It should be understood that although the above preparation scheme only provides the preparation of compounds BIO and CBIO, those skilled in the art can prepare the compounds of formula II and formula I of the present invention by changing the groups of the reactants.

[0062] The present invention will be specifically described below through examples. All chemical reagents and solvents used in the examples were purchased from commercial sources and used without further purification. Thin layer chromatography (TLC) was performed on silica gel plates. Column chromatography was performed using 200-300 mesh silica gel (Hailang, Qingdao). Chemical shifts were recorded in ppm on a Bruker AV-400 spectrometer (in deuterochloroform with MeSi as internal standard) 1 H and 13 C NMR.

Embodiment 1

[0063] Embodiment 1: the synthetic route of fluorescent polarization probe BIO

[0064] (1) Preparation of N-butyl-4-bromo-1,8-naphthalimide (BIO1)

[0065]

[0066] The synthesis of BIO1 was improved on the basis of literature (Lou, Z.R.; Li, P.; Pan, Q.; Han, K.L. Chem Commun 2013, 49, 2445). The specific operation is: weigh 7-9g of 4-bromo-1,8-naphthalic anhydride and dissolve it in 100-200mL of absolute ethanol (EtOH), add 70-80mL of n-butylamine under nitrogen protection conditions, Then the above reaction mixture was heated to 80°C, refluxed, stirred for 4h, followed by thin-layer chromatography (TLC), and after the reaction, the mixture was cooled to room temperature, filtered with suction, washed with ethanol three times, and the filtrate was taken, and the solvent was evaporated by rotary evaporation. Evaporate to dryness, column chromatography silica gel separation, dichloromethane: methanol = 50:1 as a developing solvent, to obtain a light yellow solid product B...

Embodiment 2

[0083] Implementation 2: Fluorescence Polarization Probe BIO Selectivity

[0084] Hela cells were cultured in DMEM medium containing 10% fetal bovine serum, 5% CO2, and 37°C. Dissolve the fluorescent polarization probe BIO and the control probe CBIO in water and DMSO respectively to make 1.2mg mL -1 probe master solution and 10 -3 M's control probe stock solution. Add the probe master solution and 2 mL of PBS buffer (pH=7.4) to the cuvette, so that the final concentration of the probe is 14.56 μg mL -1 As the blank control group of the probe, use the fluorescence polarization module of the fluorescence spectrometer to measure the fluorescence polarization value (Ex.Slit Width=10nm, Em.Slit Width=10nm, Slow), the emission wavelength range is 540-560nm, and the excitation wavelength is 450nm . After the measurement of the probe blank control group, the cuvette was taken out and Hela cells were added to make the cell concentration in the system 10 5 cell mL -1 As the probe ...

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Abstract

The invention relates to a construction method and an application of a micro-molecular fluorescence probe-based novel fluorescence polarization sensor, and concretely relates to a compound represented by formula I, a preparation method thereof, a detection composition containing the compound, and a detection kit. The invention also relates to a method using the compound represented by the formula I to detect the content of a CD44 protein in a biological system.

Description

technical field [0001] The invention belongs to the technical field of protein biological detection, specifically relates to CD44 protein content measurement in biological systems and tumor cell quantitative detection technology, and is a high-sensitivity fluorescence polarization measurement probe that can be used for CD44 protein fluorescence imaging in living cells and tumor cells. Background technique [0002] CD44 protein is a cell surface transmembrane glycoprotein with a very wide distribution of multi-molecular forms, which belongs to the family of adhesion molecules and has a series of physiological functions, including: (1) as a guiding receptor, regulating lymphocytes in blood and lymph (2) play a role in the autolysis of lymphocytes and the activation of isolated lymphocytes; (3) promote the interaction between fibroblasts and lymphocytes and extracellular matrix components such as hyaline Adhesion of uric acid, chondroitin sulfate, cellulose, glycogen, etc.; (4)...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N21/64C08B37/08
Inventor 黄楚森贾倜贾能勤付聪颖
Owner SHANGHAI NORMAL UNIVERSITY
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