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Dead/living cell distinguishing fluorescence probe and synthesis method and application thereof

A technology of fluorescent probes and synthesis methods, applied in the direction of fluorescence/phosphorescence, chemical instruments and methods, analytical materials, etc., can solve the problems of lack of distinguishing detection, etc., and achieve the effect of high yield and simple synthesis method

Inactive Publication Date: 2018-08-07
UNIV OF JINAN
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Aiming at the current lack of fluorescent probes for distinguishing dead / living cells, the present invention provides a fluorescent probe for distinguishing dead / living cells with variable fluorescent colors

Method used

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  • Dead/living cell distinguishing fluorescence probe and synthesis method and application thereof
  • Dead/living cell distinguishing fluorescence probe and synthesis method and application thereof
  • Dead/living cell distinguishing fluorescence probe and synthesis method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0032] Example 1 Synthesis of fluorescent probes.

[0033] (1) Synthesis of 1,2-dimethyl-quinoline iodide salt (compound 1):

[0034] Add 5-10 mL of ethanol to the round bottom flask, then add 1.1-1.4 mL of 2-methylquinoline, add 0.5-1.2 mL of iodomethane solution, heat to 40-60 degrees for 24-48 hours, and the reaction is complete After cooling the reaction system to room temperature, a solid precipitated out, filtered and washed with ethanol to obtain 1,2-dimethyl-quinoline iodide salt (compound 1), with a yield of 92%. Its H NMR spectrum is as figure 1 shown. 1 H NMR (400 MHz, DMSO- d 6 ) δ 9.12 (d, J = 8.5 Hz, 1H), 8.60 (d, J = 9.0 Hz, 1H), 8.41 (dd, J = 8.2, 1.6 Hz, 1H), 8.24 (ddd, J = 8.8, 7.0, 1.6Hz, 1H), 8.14 (d, J = 8.5 Hz, 1H), 8.00 (t, J = 7.6 Hz, 1H), 4.45 (s, 3H),3.09 (s, 3H). 13 C NMR (101 MHz, DMSO- d 6 ) δ (ppm): 161.63, 145.85, 139.65, 135.52, 130.77, 129.46, 128.24, 125.62, 119.49, 40.66, 40.45, 40.36, 40.24, 40.03, 2, 39.63, 39.62

[0035...

Embodiment 2

[0037] Example 2 Cell imaging of fluorescent probe MNQI.

[0038](1) Cell culture, treatment and staining:

[0039] Set the density to 3 x 10 5 cells / mL of HeLa cells were inoculated into sterile 35 mm imaging culture dishes in CO 2 Incubator (at 37°C, 5% CO 2 ) for more than 12 hours to allow the cells to adhere to the wall. Adherent cells were treated with 4% paraformaldehyde for 30 min to obtain dead cell samples. The DMSO solution of the probe obtained in Example 1 was prepared at a concentration of 2.5 mM as the mother solution, and the mother solution was added to the dead and living cell culture dish so that the final concentration was 5 μM. Continue to culture under the same conditions for 1 h, then aspirate the cell culture medium, wash the cells with the medium for 3 times, and then perform the cell imaging experiment.

[0040] (2) Confocal microscope imaging:

[0041] With 488 nm as the excitation wavelength, the collection wavelength of the green light chann...

Embodiment 3

[0042] Example 3 Probe MNQI co-localizes with commercial probes.

[0043] In order to further confirm the coloring position of the probe MNQI in dead and living cells, the commercial mitochondrial dye Mitochondrial Deep Red (MTDR) and the commercial nuclear dye (Hoechst33342) were used to perform colocalization staining imaging with MNQI in living cells and dead cells, respectively.

[0044] In the cell colocalization experiment, the cells were first stained with 200 nM MTDR for 30 min, and then 5 μM MNQI was added to stain the cells for 60 min, then the cell culture medium was aspirated, and the cells were washed with the medium for 3 times for cell imaging. Use 488 nm as the excitation wavelength, collect the fluorescence at 570-620 nm to collect the fluorescence signal of MNQI; use 633 nm as the excitation wavelength, collect the fluorescence at 663-738 nm to collect the fluorescence signal of MTDR. Get the fluorescence picture as Figure 6 The first column shows, from lef...

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Abstract

The invention provides a target and fluorescence color variable fluorescence probe and application thereof in distinguishing of imaging dead and living cells. A chemical name of the fluorescence probeis 2-(6-methoxy-6-naphthylvinyl)-N-methyl-quinoline iodate. The fluorescence probe is synthesized by steps: using 2-methylquinoline and iodomethane to synthesize 1,2-dimethylquinoline iodate; subjecting the 1,2-dimethylquinoline iodate and 6-methoxy-2-naphthaldehyde to condensation reaction at the room temperature to generate a product by taking pyrrolidine as a catalyst. The synthesis method ofthe probe is simple, dead and living cells can be distinguished and marked by two fluorescence colors and two dyeing positions, and a promising application prospect is achieved.

Description

technical field [0001] The invention relates to a fluorescent probe for distinguishing between living and dead cells with double targets and variable fluorescent color, as well as a synthesis method and application, and belongs to the field of small organic molecule fluorescent probes. Background technique [0002] Distinguishing detection of dead and living cells is of great significance in biology, medical science and related fields. In biology, distinguishing and detecting dead and living cells is an important tool for studying the process of apoptosis; in the field of medicine, distinguishing and detecting dead and alive cells and counting cell survival rate are the most direct methods to confirm drug efficacy and cytotoxicity. Therefore, reagents that can distinguish between living and dead cells are important research tools in the field of life sciences, can promote the development of life sciences, and have broad commercialization prospects. [0003] Until now, the d...

Claims

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Application Information

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IPC IPC(8): C07D215/14C09K11/06G01N21/64
CPCC07D215/14C09K11/06C09K2211/1029G01N21/6486
Inventor 林伟英田明刚孙洁
Owner UNIV OF JINAN
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