A membrane-permeable dye with a large two-photon fluorescence active cross-section and its application

A two-photon fluorescence and permeability technology, applied in the direction of luminescent materials, fluorescence/phosphorescence, styrene-based dyes, etc., can solve the problems that restrict two-photon three-dimensional imaging technology for cell biological imaging and observation, and achieve excellent cell membrane permeability , excellent low toxicity, good biocompatibility

Inactive Publication Date: 2017-03-22
SHANDONG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

This largely restricts the use of two-photon 3D imaging technology for cell biological imaging and observation.

Method used

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  • A membrane-permeable dye with a large two-photon fluorescence active cross-section and its application
  • A membrane-permeable dye with a large two-photon fluorescence active cross-section and its application
  • A membrane-permeable dye with a large two-photon fluorescence active cross-section and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0027] Synthesis of 4,4-2 Formyl Triphenylamine

[0028] 20ml of DMF was added to the single neck flask. Under stirring in an ice-water bath, 20 ml of phosphorus oxychloride was slowly added dropwise to the above solution, and stirred at room temperature for 1 h to obtain a yellow turbid liquid. 5 g (20.4 mmol) of triphenylamine was weighed, dissolved in 10 ml of chloroform, and then gradually added to the above-mentioned stirred solution, and the reaction was heated under reflux for 10 hours. The reaction solution was poured into 1000 mL of water, and CH 2 Cl 2 extraction. The organic layer was washed with saturated sodium chloride solution, anhydrous MgSO 4 Dry, filter and evaporate the solvent. The crude product was separated by column chromatography, and petroleum ether / ethyl acetate (10:1) was used as the eluent to obtain a light yellow solid, which was 4,4-2 formyl triphenylamine. Yield: 55%.

[0029] 1 H NMR (400MHz, d6-DMSO): δ (ppm) 9.88 (s, 2H), 7.85 (d, J=8....

Embodiment 2

[0037] 4-[(E)-2-(1H-benzimidazol-2-yl)vinyl]-N-{4-[(E)-2-(1H-benzimidazol-2-yl)vinyl] Synthesis of Phenyl}-N-phenylaniline

[0038] The synthesis technique is the same as in Example 1, and 4-[(E)-2-(1H-benzimidazol-2-yl)vinyl]-N-{4-[(E)-2-(1H-benzoyl) is obtained by synthesis Imidazol-2-yl)vinyl]phenyl}-N-phenylaniline, yield: 17%.

[0039] 1H NMR (400MHz, d6-DMSO): δ (ppm): 12.55 (s, 2H), 7.65 (m, J=7.67Hz, 6H), 7.55 (d, J=7.64Hz, 4H), 7.41 (t, J=7.84Hz, 2H), 7.16 (t, J=7.76Hz, 5H), 7.11 (d, J=7.68Hz, 4H), 7.07 (d, J=8.24Hz, 4H).

Embodiment 3

[0041] RBL-2H3 and Hela cell culture

[0042] RBL-2H3 or HeLa cell lines were adherently cultured in medium containing 10% fetal bovine serum at 37°C, 5% CO 2 Cultured in a saturated humidity incubator, and the medium was changed every 2 to 3 days for passage. After the cells grow to the logarithmic phase, the slicing culture: ① Soak the coverslip in absolute ethanol for 30 minutes, dry it in an alcohol lamp and put it into a disposable 35mm petri dish; ② Wash the cells in the 100ml cell flask with PBS Three times, digest with 1 ml of 0.25% trypsin for 3-5 minutes, carefully pour out the medium, add a small amount of fresh medium and pipet evenly, after counting the cells, leave the cells of the appropriate density, and add the medium to the desired volume ( Control cells at a final concentration of 1x10 5 ), inoculated into a Petri dish containing a coverslip, placed in CO 2 Cultured in an incubator to allow the cells to grow on the sheet.

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Abstract

The invention discloses membrane permeability dye with a large two-photon fluorescence active cross section. The dye adopts triphenylamine heterocyclic chemical compounds, and the chemical structure of the dye is shown in the formula (I). The invention further discloses an application of the dye in displaying two-photon imaging of cytoplasm in a living cell. Experiments show that the dye has characteristics of larger two-photon fluorescence active absorption cross section, excellent cell membrane permeability, low toxicity and the like, also has the characteristics of wide application range, low price and good bio-compatibility with known probe DAPI (4',6-diamidino-2-phenylindole) and has wide application prospect in the field of laser excitation fluorescence biomarkers.

Description

technical field [0001] The invention relates to a two-photon dye and its application, in particular to a membrane-permeable dye with a large two-photon fluorescent active cross section and its application. Background technique [0002] The development of two-photon microscopy has revolutionized the observation of organs and tissues, especially thick or highly scattering specimens. Compared with single-photon laser confocal microscopy, two-photon microscopy has features such as near-infrared excitation, avoidance of fluorescence bleaching and phototoxicity, high resolution, reduced tissue absorption, and reduced tissue autofluorescence interference. However, if the two-photon absorption cross-section of the fluorescent probe used is small (similar to the endogenous luminescent substance of the biological sample), the advantages of two-photon microscopy will be greatly reduced. Because the one-photon and two-photon absorption of dyes obey different selection laws, a good one-...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C09B23/14C09K11/06C07D277/64C07D235/14G01N21/64
Inventor 于晓强冯瑞卿孙渝明李学晨
Owner SHANDONG UNIV
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