Fluorescent dye for marking lysosome as well as synthesis method and application of fluorescent dye
A fluorescent dye and lysosome technology, applied in the field of fluorescent dye and fluorescent imaging, can solve the problems of small Stokes shift, poor structural stability, complex synthesis route, etc., achieve large Stokes shift and simple preparation method , The effect of high fluorescence quantum yield
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Embodiment 1
[0020] Synthesis of fluorescent dye molecule Lyso-DNBN with the structure of the compound of formula I:
[0021]
[0022] 6-Bromo-N,N-dimethylnaphthalene-2-amine (100mg, 0.40mmol), palladium acetate (15mg, 0.07mmol) and triphenylphosphine (53mg, 0.20mmol) were dissolved in N,N-di In a mixed solvent of methylformamide (25 mL) and triethylamine (20 mL). After the temperature of the reaction solution was raised to 45° C. for 0.5 hour, 4-vinylbenzonitrile (62 μL, 0.54 mmol) was added. The reaction solution was heated to 110° C. for 8 hours. The solvent was distilled off under reduced pressure, and the obtained crude product was separated and purified by chromatographic column, the silica gel used was 300-400 mesh, and the eluent was a mixed solvent of petroleum ether and ethyl acetate with a volume ratio of 10:1. The solvent was removed by rotary evaporation to obtain 83 mg of a yellow solid product with a yield of 71%.
[0023] The structure of the product was characterized...
Embodiment 2
[0026] Optical properties such as ultraviolet absorption and fluorescence emission of the fluorescent dye molecule Lyso-DNBN prepared in Example 1 were studied.
[0027] The fluorescent dye molecule Lyso-DNBN was dissolved in DMSO solution with a concentration of 10 -5 M solution to be tested. Through ultraviolet absorption and fluorescence tests, it is found that the maximum absorption wavelength of the fluorescent dye molecule Lyso-DNBN in DMSO solution is 401nm, the maximum fluorescence emission wavelength is 592nm, and the Stokes shift can reach 191nm, as figure 1 shown. The fluorescent quantum yield of Lyso-DNBN in DMSO solution is 73.9%.
Embodiment 3
[0029] Colocalization of the fluorescent dye molecule Lyso-DNBN prepared in Example 1 and lysosome blue in SKBR-3 cells
[0030] SKBR-3 cells were co-incubated with the fluorescent dye molecule Lyso-DNBN (2 μM) prepared in Example 1 and the commercial lysosome labeling dye LysoBrite Blue (2 μM), and fluorescent confocal imaging was performed after 10 minutes. Fluorescent staining in the blue and yellow channels was monitored by confocal microscopy. like figure 2 As shown, the excitation wavelength of the blue channel is 405 nm, and the fluorescence signal of 430-450 nm is collected, and the excitation wavelength of the yellow channel is 405 nm, and the fluorescence signal of 490-530 nm is collected. Comparing the fluorescence images of the two channels, it is found that the lysosome blue fluorescence signal of the blue channel and the fluorescence signal of the fluorescent dye molecule Lyso-DNBN of the yellow channel have a high degree of overlap, and the overlap correlation...
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