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Disposable and rapid octadecyl-chain-containing targeting mitochondrion fluorescent probe and application thereof

An octadecyl chain and fluorescent probe technology, which is applied in the direction of fluorescence/phosphorescence, luminescent materials, and material analysis through optical means, can solve the problem of reducing the signal-to-noise ratio of pictures, long dyeing time of probes, and inability to wash tissues and other issues, to achieve the effect of broad application prospects, intuitive biological detection reagents, and good biocompatibility

Active Publication Date: 2017-04-26
SHANDONG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, in the process of tissue imaging, the inside of the tissue cannot be washed, resulting in free probes that will reduce the signal-to-noise ratio of the image
In addition, for tissue staining, the long staining time of the probe is also a disadvantage

Method used

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  • Disposable and rapid octadecyl-chain-containing targeting mitochondrion fluorescent probe and application thereof
  • Disposable and rapid octadecyl-chain-containing targeting mitochondrion fluorescent probe and application thereof
  • Disposable and rapid octadecyl-chain-containing targeting mitochondrion fluorescent probe and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0027] Synthesis of Mito-19

[0028] Dissolve 9-octadecyl-9H-carbazole-3-carbaldehyde (0.447g, 1mmol) and 1,3,3-trimethyl-3H-indole-1-iodide (0.301g, 1mmol) in Add 4 drops of piperidine to 15 mL of methanol, and the solution turns red rapidly. Heated to reflux overnight, cooled and poured the mixture into petroleum ether, a red solid was precipitated, and ethanol was used as eluent to obtain small red crystals. Yield 38%.

[0029] 1 H NMR (400MHz, CDCl 3 )δ (ppm): 8.99 (s, 1H), 8.37-8.48 (m, 3H), 7.97 (d, J = 15.68Hz, 1H), 7.48-7.55 (m, 6H), 7.43 (d, J = 8.12 Hz, 1H), 7.36(t, J=7.42Hz, 1H), 4.45(s, 3H), 4.30(t, J=7.26Hz, 2H), 1.87-1.89(m, 6H), 1.24-1.25(m ,32H),0.87(t,J=6.82Hz,3H). 13C NMR(400MHz,CDCl3),δ(ppm):181.08,156.95,144.60,142.49,141.59,130.38,129.41,128.82,127.05,126.52,125.45,124.32,123.09,122.39,122.20,121.14,113.83,110.16,109.51 ,109.08,77.48,77.06,76.64,51.72,43.63,36.41,31.92,29.70,29.65,29.61,29.57,29.49,29.36,28.98,27.46,27.24,22.68,14.13.HRMS(3.45,C0)=6...

Embodiment 2

[0031] Culture of Immortalized Cells (SiHa and HeLa) and Normal Cells (Rat Astrocytes)

[0032] All cell lines were stored at 37°C, 5% CO 2 cultured in a saturated humidity incubator. SiHa and HeLa cell lines were cultured adherently in H-DMEM medium containing 10% fetal bovine serum (containing 1% double antibody). Rat astrocytes were cultured adherently in F-12 medium containing 10% fetal bovine serum. After the cells grow to the logarithmic phase, culture the slices: ① Soak the coverslips in absolute ethanol for 30 minutes, dry them with an alcohol lamp, and place them in a disposable 35mm culture dish; ② Wash the cells in the 100mL cell bottle with PBS. Three times, digest with 1mL 0.25% trypsin for 3-5min, pour out the medium carefully, add a small amount of fresh medium and pipette evenly, after counting the cells, leave cells with a suitable density, add the medium to the required volume (control The final concentration of cells was 1 × 10 4 ), inoculated into a pet...

Embodiment 3

[0034] Mito-19 staining rat astrocytes and MTG counterstaining experiments

[0035] Wash the inoculated cell slide three times with PBS, stain the cells with 5 μM Mito-19, and store in CO 2 30min in the incubator. Aspirate the culture medium and wash it three times with PBS to wash off the unbound excess dye, stain the cells with 0.2 μM MTG, and store in CO 2 30min in the incubator. The stained cell slides were taken out, the unbound excess dye solution was washed away, and the cell growth side was covered on the glass slide, and the stained parts, fluorescence distribution and brightness changes of the cells were observed under a laser scanning confocal fluorescence microscope. The results found that , MTG and Mito-19 were similar in intracellular distribution area, and the average colocalization rate of 2 astrocytes was calculated to be 0.98. Therefore, it is confirmed that the probe Mito-19 of the present invention can specifically image mitochondria.

[0036] see resul...

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PUM

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Abstract

The invention discloses a disposable and rapid octadecyl-chain-containing targeting mitochondrion fluorescent probe. The fluorescent probe is a carbazole pseudo-indole salt compound of a structure shown in a formula (I). The compound can achieve disposable and rapid imaging of mitochondria in living cells and tissue. Particularly, in rat skeletal muscle tissue, the probe can achieve clear imaging of mitochondrion reticular structures of four different mitochondria, and can give a three-dimensional high-definition image of distribution of mitochondrion in the tissue. The probe can track dynamic changes of the mitochondrial morphology in cells in real time. Compared with a mitochondrion fluorescent probe with similar functions, the fluorescent probe has the advantages of being low in cost, good in light stability and good in biocompatibility. The probe also has the high mitochondria positioning capability and hypotoxicity, has the good biocompatibility with Hochest 33342, MTG and MTR, and has the potential application value in the field of laser induced fluorescence biomarkers.

Description

technical field [0001] The present invention relates to a mitochondria-targeted probe and its application, in particular to an octadecyl chain-containing fluorescent probe that is free from washing and rapidly targets mitochondria and its application in live cell and tissue imaging. Background technique [0002] To obtain more intrinsic biological information, it is important to image organelles in situ in intact living tissues. In living tissues, the interaction of cells with neighboring cells and the extracellular matrix forms a three-dimensional (3D) network to maintain tissue specificity and homeostasis. For a long time in the past, many research results of cell biology were obtained from cells cultured in vitro. But recent studies have shown that there are indeed phenotypic differences between cultured cells and real tissues. Therefore, the development of tools that enable in situ studies in tissues is highly desirable. Fluorescent imaging technology based on biolumi...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07D209/86C09K11/06G01N21/64
CPCC07D209/86C09K11/06C09K2211/1029G01N21/6486
Inventor 于晓强张若瑶孙渝明张革
Owner SHANDONG UNIV
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