Fluorescent probe and application thereof in detection of autolysosome

A technology of fluorescent probes and stereoisomers, applied in the field of analytical chemistry, can solve problems such as lack, and achieve the effects of good biocompatibility, low cytotoxicity, and easy detection and operation.

Active Publication Date: 2020-04-21
INST OF CHEM CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

There are several fluorescent probes for labeling lysosomes on the market, such as LysoTracker Red...

Method used

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  • Fluorescent probe and application thereof in detection of autolysosome
  • Fluorescent probe and application thereof in detection of autolysosome
  • Fluorescent probe and application thereof in detection of autolysosome

Examples

Experimental program
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Embodiment 1

[0041] 1. The synthesis methods of compounds (1)-(5) are as follows, wherein the reaction ratios and purification methods adopt conventional ratios or conventional purification methods in the art. In addition, the inventors have confirmed that the structure of the above-mentioned compounds is correct by analyzing the hydrogen spectrum, carbon spectrum and / or mass spectrum data of each compound, wherein the mass spectrum and mass spectrum of compound (2) 1 H-NMR spectra were referred to Figure 8 and Figure 9 shown.

[0042]

Embodiment 2

[0044] Compounds of formulas (1)-(5) have similar properties, and the following mainly describes their properties by taking the compounds of formulas (1)-(3) as fluorescent probes (1)-(3) respectively as examples.

[0045] Utilize the fluorescent probe (1) of the embodiment of the present invention to carry out cytotoxicity experiment, specifically as follows:

[0046]

[0047] (1) Dissolving the fluorescent probe (1) with a small amount of methanol;

[0048] (2) The cultured HeLa and MCF-7 cells were added different concentrations of probe (1) solutions and continued to culture for 24 hours;

[0049] (3) Add 10% MTT solution after sucking up the culture medium, and continue culturing for 4 hours;

[0050] (4) After the culture medium was blotted dry, DMSO was added to dissolve it, and the absorbance at 559 nm was measured by a microplate reader. Take the absorbance value at 559 nm as the ordinate, and the concentration value of the probe (1) as the abscissa, and draw a g...

Embodiment 3

[0052] Utilize the fluorescent probe (1) of the embodiment of the present invention to assemble in an aqueous solution containing potassium ions to form supramolecular J-aggregates, as follows:

[0053]

[0054] (1) Dissolving the fluorescent probe (1) with a small amount of methanol;

[0055] (2) Dilute the fluorescent probe solution in step (1) with water containing different concentrations of potassium ions, and configure them into aqueous solutions containing the fluorescent probe (1) at a concentration of 4 μM;

[0056] (3) Utilize ultraviolet absorption spectrometer to detect the absorption spectrum of step (2) fluorescent probe (1) aqueous solution, the result is as follows figure 2 As shown, with the increase of potassium ion concentration, a new absorption peak was generated at 650nm, indicating that probe (1) formed supramolecular J-aggregates.

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Abstract

The invention discloses a fluorescent probe and new application of the fluorescent probe in detection of autolysosome of living cells. The fluorescent probe contains a compound having a structure represented by the following formula or a stereoisomer thereof and countra-ions. The fluorescent probe can form a supramolecular aggregate form through self-assembly in a solution system containing metalsalt ions or an intracellular physiological environment; the fluorescent probe has two excitation wavelengths of 530-570nm and 600-650nm, can realize real-time observation of the autophagy process ofa living cell sample, has the advantages of simple dyeing, low cytotoxicity and small damage to a biological sample, and is not influenced by the pH value in cells.

Description

technical field [0001] The present invention relates to the field of analytical chemistry, in particular, to fluorescent probes and their use in detecting autophagy lysosomes, more specifically, to fluorescent probes and their use in detecting autophagy lysosomes and determining cell A method for the presence or absence of autophagic lysosomes. Background technique [0002] Lysosome is a very important organelle with a diameter of about 200-500nm, which exists in almost all eukaryotic cells. There are many enzymes and proteins in lysosomes, including acid hydrolase, trypsin and cathepsin, which are digestive organs in cells, which can decompose various proteins and other macromolecules, and have the function of dissolving or digesting. Autophagy lysosomes, also known as cytolysosomes, are formed by the fusion of primary lysosomes and vesicles containing endogenous substances from autophagy, or lysosomes engulfing the cytoplasm. Acts as a "scavenger" for the normal pathway ...

Claims

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Application Information

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IPC IPC(8): C07D277/84C09K11/06G01N21/64
CPCC07D277/84C09K11/06G01N21/6428C09K2211/1037C09K2211/1007G01N2021/6439Y02A50/30
Inventor 孙红霞唐亚林
Owner INST OF CHEM CHINESE ACAD OF SCI
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