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Fluorescent probes for labelling and/or detecting lipid droplets in cells as well as preparation and applications of fluorescent probes

A fluorescent probe and quasi-labeling technology, applied in the direction of fluorescence/phosphorescence, luminescent materials, measuring devices, etc., can solve the problems of unfavorable long-term fluorescence imaging analysis, easy fluorescence quenching, fluorescence instability, etc., and achieve convenient post-processing , good light stability, cheap and easy-to-obtain raw materials

Inactive Publication Date: 2018-05-25
DALIAN INST OF CHEM PHYSICS CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Nile red is used to label lipid droplets, its specificity is not high, and it can stain most structures in the cell, so the background is strong, and it is difficult to distinguish lipid droplets from other structures in the cell
Fluoroboron fluorescent probes are not conducive to long-term fluorescence imaging analysis due to their small Stokes shift and unstable fluorescence, which is prone to fluorescence quenching.

Method used

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  • Fluorescent probes for labelling and/or detecting lipid droplets in cells as well as preparation and applications of fluorescent probes
  • Fluorescent probes for labelling and/or detecting lipid droplets in cells as well as preparation and applications of fluorescent probes
  • Fluorescent probes for labelling and/or detecting lipid droplets in cells as well as preparation and applications of fluorescent probes

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0039] Example 1: The preparation of the fluorescent probe N-dodecyl-4-azetidine-1,8-naphthalimide, the basic synthesis process is as follows:

[0040] (1) Synthesis of N-dodecyl-4-bromo-1,8-naphthalimide:

[0041] In a 250mL single-necked bottle, 2.27g (10mmol) of 4-bromo-1,8-naphthalene anhydride and 2.78g (15mmol) of dodecylamine were dissolved in 100mL of ethanol, and refluxed at 90°C for 12 hours. After the heating was stopped, the precipitated solid was filtered and washed three times with 20 mL of ethanol to obtain 3.2 g of N-dodecyl-4-bromo-1,8-naphthalimide with a yield of 72%.

[0042] (2) Synthesis of N-dodecyl-4-azetidine-1,8-naphthalimide:

[0043] 0.44g (1mmol) N-dodecyl-4-bromo-1,8-naphthalimide, 0.1mL (1.5mmol) azetidine The alkane was dissolved, and reacted at 140° C. for 8 hours under nitrogen protection. After spin-drying the solvent, after separation by 200-300 mesh silica gel column chromatography at 25°C, spin-dry to obtain 280 mg of a red waxy solid t...

Embodiment 2

[0046] Embodiment 2: the mensuration of the ultraviolet-visible absorption spectrum of fluorescent probe:

[0047] The probe synthesized in Example 1 was prepared at a concentration of 10 μM with 20 mM HEPES solution (pH=7.4). Scan absorption in the 250nm-600nm band with a UV-visible spectrophotometer to obtain Figure 5 Mid-UV-Vis absorption spectrum.

Embodiment 3

[0048] Example 3: Determination of the fluorescence emission spectrum of the fluorescent probe.

[0049] The probe synthesized in Example 1 was prepared at a concentration of 10 μM with 20 mM HEPES solution (pH=7.4). Scanned with a fluorescence spectrometer under 450nm excitation Image 6Fluorescence emission spectra in .

[0050] Example 4 The fluorescent probe probe prepared in Example 1 labeled cells

[0051] Figure 7 The medium cell is colon cancer cell HT-29, and the probe used is synthesized in Example 1 at a concentration of 5 μM. HT-29 cells were inoculated in confocal small dishes at a concentration of 20,000 cells / dish, and cultured in 1640 medium for 3 days. Aspirate the medium, wash it twice with PBS buffer solution, add 2mL of fresh 1640 medium, and then add the probe solution synthesized in Example 1 at a concentration of 2mM, so that the concentration in the medium is 5μM, and in a CO2 incubator at 37°C After incubating for 10 min, the medium was sucked of...

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Abstract

The invention belongs to the field of biological analysis and detection and relates to fluorescent probes for labelling and / or detecting lipid droplets in cells as well as preparation and applicationsof the fluorescent probes. The probes are synthesized with 4-bromo-1,8-naphthalic anhydride as an initial material and take 1,8-naphthalimide as a fluorophore, a C12 alkyl long chain as a specific lipid droplet positioning group and an N-containing group on the 4th site of a naphthalene ring as a fluorescence enhanced and stabilized group. The probes are simple to prepare and have high yield. Compared with existing Nile red and boron dipyrromethene lipid droplet fluorescent probes, the probes have high light stability and large stokes shift and are more suitable for fluorescence imaging. Theprobes have very important application value in cell labelling fluorescent imaging and biomedical fields.

Description

technical field [0001] The invention belongs to the field of biological analysis and detection, and in particular relates to a fluorescent probe for marking and / or detecting lipid droplets in cells and its preparation method and application. Background technique [0002] Lipid droplet is a complex and dynamically changing multifunctional organelle, which consists of a monolayer of phospholipids and neutral lipids to form a hydrophobic core, and a variety of proteins are distributed on the surface. For a long time, lipid droplets have been considered as an inert inclusion used to store energy, so they have not attracted the attention of researchers, but the latest research shows that lipid droplets are involved in transmembrane transport, signal transduction and special circumstances Energy supply is closely related. Lipid droplets widely exist in almost all types of cells. Studies have shown that changes in the content of lipid droplets in cells are closely related to metab...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07D221/14C09K11/06G01N21/64
CPCC07D221/14C09K11/06C09K2211/1029G01N21/6486
Inventor 徐兆超邓霏
Owner DALIAN INST OF CHEM PHYSICS CHINESE ACAD OF SCI
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