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Self-flashing fluorescent dye for long-time super-resolution fluorescence imaging of lysosome as well as synthesis method and application of self-flashing fluorescent dye

A technology of super-resolution fluorescence and fluorescent dyes, applied in the direction of fluorescence/phosphorescence, azo dyes, organic dyes, etc., can solve the problem of difficult to realize the transition to dark state, and achieve the effect of simple synthesis method

Pending Publication Date: 2022-04-01
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

However, there are still few self-blinking super-resolution fluorescent dyes targeting lysosomes, mainly because most of the dyes exist in the fluorescent state under the acidic pH condition of the lysosome, and it is difficult to achieve the transition to the dark state.

Method used

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  • Self-flashing fluorescent dye for long-time super-resolution fluorescence imaging of lysosome as well as synthesis method and application of self-flashing fluorescent dye
  • Self-flashing fluorescent dye for long-time super-resolution fluorescence imaging of lysosome as well as synthesis method and application of self-flashing fluorescent dye
  • Self-flashing fluorescent dye for long-time super-resolution fluorescence imaging of lysosome as well as synthesis method and application of self-flashing fluorescent dye

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0033] Synthesis of Dye Lyso-532

[0034]

[0035] Rho19 (100mg, 0.22mmol) was placed in 20mL of 1,2-dichloroethane, then 1.0mL of phosphorus oxychloride was added to the reaction solution, and reacted at 80°C for 4h. The reaction solution was cooled and removed under reduced pressure to obtain a purple crude product. The crude product was dissolved in 20 mL of acetonitrile and 0.5 mL of triethylamine was added followed by 2-amino-6-picoline (400 mg, 2.16 mmol). Reaction at 80°C for 10h. The solvent was removed under reduced pressure, and basic alumina column chromatography (developing solvent: dichloromethane:methanol=400:1; volume ratio) gave 50 mg of white solid with a yield of 45%.

[0036] Its high-resolution mass spectrometry data are as follows:

[0037] HRMS(ESI)m / z[M+H] + : Calculated: 505.2604, Experimented: 505.2652.

[0038] Its NMR spectrum is as follows figure 1 As shown, the specific data are as follows:

[0039] 1 H NMR (400MHz, CDCl 3 )δ8.23(d, J=8...

Embodiment 2

[0046] Synthesis of Dye LysoNH-532

[0047]

[0048] Rho19 (300mg, 0.67mmol) was placed in 30mL of 1,2-dichloroethane, and then 2.0mL of phosphorus oxychloride was added to the reaction solution, and reacted at 80°C for 8h. The reaction solution was cooled and removed under reduced pressure to obtain a purple crude product. The crude product was dissolved in 30 mL of acetonitrile and 0.5 mL of triethylamine was added followed by 2-amino-5-methylaminocarbonylpyridine (900 mg, 5.95 mmol). Reaction at 80°C for 24h. The solvent was removed under reduced pressure, and basic alumina column chromatography (developing solvent: dichloromethane:methanol=300:1; volume ratio) gave 93 mg of white solid with a yield of 25%.

[0049] Its high-resolution mass spectrometry data are as follows:

[0050] HRMS(ESI)m / z[M+H] + : Calculated value: 548.2662, Experimental value: 548.2671.

[0051] The specific data of its NMR spectrum are as follows:

[0052] 1 H NMR (400MHz, CDCl 3 )δ8.22(...

Embodiment 3

[0055] Synthesis of Dye LysoH-532

[0056]

[0057] Rho19 (100mg, 0.22mmol) was placed in 15mL of 1,2-dichloroethane, then 0.5mL of phosphorus oxychloride was added to the reaction solution, and reacted at 80°C for 4h. The reaction solution was cooled and removed under reduced pressure to obtain a purple crude product. The crude product was dissolved in 15 mL of acetonitrile and 0.5 mL of triethylamine was added followed by 2-aminopyridine (500 mg, 5.32 mmol). Reaction at 80°C for 12h. The solvent was removed under reduced pressure, and basic alumina column chromatography (developing solvent: dichloromethane:methanol=400:1; volume ratio) gave 74 mg of a white solid with a yield of 67%.

[0058] Its high-resolution mass spectrometry data are as follows:

[0059] HRMS(ESI)m / z[M+H] + : Calculated value: 491.2447, Experimental value: 491.2442.

[0060] The specific data of its NMR spectrum are as follows:

[0061] 1 H NMR (400MHz, CDCl 3 )δ8.23(d,J=8.1Hz,1H),8.04–8.00(m...

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Abstract

The invention relates to a self-flashing fluorescent dye for long-time super-resolution fluorescence imaging of lysosome as well as a synthesis method and application thereof, and the dye is structurally characterized in that ring locking is performed through a 2-aminopyridine derivative, so that the ratio of a fluorescent state to a dark state of dye rhodamine is regulated and controlled. The fluorescent dye provided by the invention has the characteristics of pH gt; and the dye exists in a dark state with a closed-loop structure in an aqueous solution with the concentration of 5.5, so that the dye only exists in an acidic lysosome in a small amount of fluorescence state, and accurate positioning of the lysosome is achieved. Besides, the dye has reciprocating change of a fluorescence state and a dark state in the lysosome, so that a self-flashing effect is achieved, dynamic and long-time super-resolution fluorescence imaging of the lysosome is realized, and the pH, distribution, size and the like of the lysosome are monitored.

Description

technical field [0001] The invention belongs to the field of super-resolution fluorescent dyes, in particular to a class of self-flickering fluorescent dyes for long-time super-resolution fluorescence imaging of lysosomes and their application in the field of fluorescence imaging. Background technique [0002] Lysosome is a single-membrane organelle containing a large number of hydrolytic enzymes, which can accept and degrade macromolecules through endocytosis, secretion, autophagy, and phagocytosis. The occurrence of these processes is often accompanied by highly dynamic processes such as lysosome fusion, division, and interaction with other organelles. In situ, real-time analysis of lysosomes in living cells is essential for lysosome-related studies. Fluorescence microscopy has become an important analytical method for real-time monitoring of lysosomes due to its unique advantages. In particular, the rapid development of super-resolution imaging technology in recent year...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C09K11/06C09B57/00C07D491/107G01N21/64
Inventor 徐兆超许宁尹文婷乔庆龙
Owner DALIAN INST OF CHEM PHYSICS CHINESE ACAD OF SCI
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