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Fluorescent probe based on fused ring oxidized thiophene and application of fluorescent probe in cell imaging

A fluorescent probe, thiophene technology, applied in the field of organic chemistry and chemical biology, to achieve the effect of simple operation, high biocompatibility and high sensitivity

Active Publication Date: 2019-05-07
SOUTH CHINA UNIV OF TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, the research on the interaction between lipid droplets and lysosomes mainly uses molecular biology, genetics and other means. Although fluorescent proteins have been used to study the interaction between lipid droplets and lysosomes recently, the use of fluorescent probes for There is little work examining the interaction between the two

Method used

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  • Fluorescent probe based on fused ring oxidized thiophene and application of fluorescent probe in cell imaging
  • Fluorescent probe based on fused ring oxidized thiophene and application of fluorescent probe in cell imaging
  • Fluorescent probe based on fused ring oxidized thiophene and application of fluorescent probe in cell imaging

Examples

Experimental program
Comparison scheme
Effect test

preparation example Construction

[0054] The preparation steps of TPA-BTTDO are:

[0055] (1) Synthesis of intermediate 1

[0056] 3,6-Dibromothieno[3,2-b]thiophene (1.49g, 5mmol), phenylboronic acid (2.44g, 20mmol), tetrakis(triphenylphosphine)palladium (0.58g, 0.5mmol) and carbonic acid Potassium (3.45g, 20mmol) was added to a 250mL two-necked bottle, nitrogen was replaced three times, and THF (80mL) and H 2 O (20mL), after the injection was completed, the reaction was refluxed at 80°C for 24h. The reaction was quenched by adding water, extracted, concentrated, powdered, separated and purified by column chromatography to obtain intermediate product 1 with a yield of 75%.

[0057] (2) Synthesis of intermediate 2

[0058] Intermediate 1 (5.84g, 20mmol) was added to a 250mL single-necked bottle, dissolved with 100mL of N,N-dimethylformamide (DMF), and N-bromosuccinimide (NBS) ( 7.12g, 40mmol) into the reaction flask, stirred overnight, quenched the reaction by adding water, extracted, concentrated, powdered...

Embodiment 1

[0066] An application of a fluorescent probe based on fused ring thiophene oxide in cell imaging, comprising the following steps:

[0067] (1) Add TPA-BTTDONPs to 1 mL of cell culture system (DMEM solution), and co-culture with HeLa cells at 37°C for 8 hours; the concentration of TPA-BTTDONPs in the cell culture system is 2.4 μg / mL;

[0068] (2) After washing with PBS for 3 times, TPA-BTTDONPs that had not interacted with the cells were removed, and then reacted with the lysosome dye Lysotracker and the lipid droplet dye BODIPY for 20 min respectively; the concentrations of Lysotracker and BODIPY in the cell culture system were 100 nM and 10 μg / mL, imaged with a confocal laser microscope.

[0069] figure 1 For the compound TPA-BTTDO and the prepared nanoparticles (TPA-BTTDONPs) ultraviolet absorption (Abs corresponding curve) and fluorescence emission spectrum (PL corresponding curve) (A), TPA-BTTDO in different proportions of tetrahydrofuran and water mixed solvent Fluores...

Embodiment 2

[0078] A fused epoxy thiophene oxide is used to trace lipid droplet movement, comprising the following steps:

[0079] (1) HeLa cells were co-cultured with a medium mixed with oleic acid (100 μM) for 6 h;

[0080] (2) After washing with PBS for 3 times, in 1 mL of cell culture system (DMEM solution), add TPA-BTTDONPs) and co-culture HeLa cells at 37°C for 8 h; the concentration of TPA-BTTDONPs in the cell culture system was 2.4 μg / mL;

[0081] (3) After washing with PBS for 3 times, the TPA-BTTDONPs that did not interact with the cells were removed, and observed under a confocal laser microscope.

[0082] Figure 5 TPA-BTTDONPs trace lipid droplet intracellular movement in Example 2, respectively 0min, 2min, 4min, 7min, 8min and 10.5min laser confocal imaging, wherein the image in the upper left corner box is indicated by a circle Enlarged view of part; where the scale bar is 5 μm.

[0083] Figure 5 Because the HeLa cells in Example 2 were induced by oleic acid, a large...

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Abstract

The invention belongs to the technical field of biochemistry, and discloses a fluorescent probe based on fused ring oxidized thiophene and application of the fluorescent probe in cell imaging. The fluorescent probe based on the fused ring oxidized thiophene is nano-particles coated with the fused ring oxidized thiophene. The fluorescent probe is applied to cell imaging, in particular to lipid droplet and lysosome imaging. The fluorescent probe can be used as a fluorescent imaging dye for lysosomes and lipid droplets, a tracer agent of mutual action between the lysosomes and the lipid droplets,and a tracer agent for lipid droplet formation and lipid droplet metabolism. The fluorescent probe shows blue-fluorescence in the lipid droplets, and shows red fluorescence in the lysosomes after entering cells, the fluorescent signal intensity and the color change are monitored, so that the high-sensitivity and high-resolution tracing of the movement of the droplets in a short time is realized;and the distribution and the metabolism conditions of the droplets are traced through long-time monitoring, and it is proved for the first time that the internal components of the droplets can be recovered by the lysosomes. The fluorescent probe is high in sensitivity and simple to operate.

Description

technical field [0001] The invention relates to the fields of organic chemistry and chemical biology, in particular to a fluorescent probe based on fused oxythiophene and its application in cell imaging, especially the application in cell lysosome and lipid droplet imaging. Background technique [0002] Lipid droplets and lysosomes play very important roles in cells. Lipid droplet is an important organelle for storing neutral lipids in cells and a storage place for energy substances and cell membrane components. In recent years, a large number of studies have found that lipid droplets are inextricably linked to many current diseases, such as obesity, type II diabetes, arteriosclerosis and even cancer. Therefore, the research on lipid droplets has attracted more and more researchers' attention. Lysosome is the "digestive organ" of the cell, which contains more than 100 kinds of hydrolytic enzymes, which can decompose the substances entering the cell from the outside, and can...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C09K11/02C09K11/06G01N21/64
Inventor 唐本忠赵祖金胡蓉陈斌秦安军
Owner SOUTH CHINA UNIV OF TECH
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