AIE nano-probe responsive to sulfatase and preparation method and application of AIE nano-probe
A technology of sulfatase and nanoprobe, which is applied in the field of biomedicine, can solve the problems of small displacement, few sulfatases, and short emission wavelength of fluorescent probes, and achieves convenient use, selective rapid detection, and simple synthesis Effect
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Embodiment 1
[0042] The preparation method of the AIE nanoprobe of sulfatase response:
[0043] (1) Synthesis of fluorophore: 2-methylquinoline (2.86g, 20mmol) and ethyl iodide (3.90g, 25mmol) were weighed and dissolved in 25mL of anhydrous acetonitrile, and stirred at 85°C for 24h under nitrogen protection. After the reaction was finished, after the reaction mixture was cooled to room temperature, there was a large amount of precipitate at the bottom of the flask, which was vacuum filtered with a Buchner funnel. The crude product was washed with cold acetonitrile without further purification. After drying overnight, compound 2 was obtained.
[0044]
[0045] (2) Compound 2 (1.72g, 10mmol) was weighed and dissolved in 10mL of dry ethanol, and then malononitrile (1g, 15mmol) and sodium ethoxide (1.02g, 15mmol) were added. The reaction was stirred at 0 °C for 0.5 h, then at room temperature for an additional 3 h. After the reaction was completed, the precipitate was filtered and washed...
Embodiment 2
[0054] Weigh 4.2 mg of the probe DQM-SULF prepared in the example and dissolve it in 10 mL of DMSO solution to prepare a standard solution with a concentration of 1 mM; similarly, take 3.4 mg of the fluorophore DQM-OH prepared in the example and dissolve it in 10 mL of DMSO solution , prepared into a standard solution with a concentration of 1mM; the sulfatase was prepared into a 100U / mL standard solution with PBS buffer solution (10Mm, pH 7.4). Take a 4mL EP tube, add different final concentrations of sulfatase enzyme solutions (0-50U / mL) to the reaction system PBS buffer (10mM, pH=7.4, 1%DMSO) containing the probe DQM-SULF (5μM). Incubate at 37°C for 30 min in a constant temperature shaker. Under the excitation of 440nm wavelength, the slit width was set as 10.0 / 10.0nm, and the fluorescence emission spectrum of the solution in the 450-800nm band was collected. Such as figure 2 As shown, the fluorescence intensity curve of the probe DQM-SULF increases with the concentrat...
Embodiment 3
[0056] The preparation steps of the probe DQM-SULF solution and the sulfatase solution involved in this embodiment are the same as those in Example 2, and other specific steps are as follows: add to the reaction system PBS buffer (10 mM, pH = 7.4, 1% DMSO) was added with a sulfatase solution at a final concentration of 50 U / mL, and co-incubated at 37° C. for 30 min in a constant temperature shaker. Under the excitation of 440nm wavelength, the slit width was set as 10.0 / 10.0nm, and the fluorescence emission spectrum of the solution in the 450-800nm band was collected. Such as image 3 As shown, the AIE fluorescence signal is amplified in the fluorophore-enzyme response compared to the fluorophore alone. It shows that the probe prepared by the present invention can release the fluorophore and realize the secondary enhancement of the fluorescence signal, effectively solving the limitations of the quenching effect caused by the short emission wavelength of the fluorescent prob...
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