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An Intracellular Detection Method Based on Dual Excitation Ratiometric Upconversion Fluorescent Probes

A detection method and nano-probe technology, applied in fluorescence/phosphorescence, material excitation analysis, measurement devices, etc., can solve problems such as inability to achieve accurate intracellular quantitative detection, and achieve the effect of improving sensitivity

Active Publication Date: 2022-02-18
FUJIAN INST OF RES ON THE STRUCTURE OF MATTER CHINESE ACAD OF SCI
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] In order to solve the problem that dye-sensitized up-conversion nanoprobes cannot achieve accurate intracellular quantitative detection in the prior art, the present invention provides an intracellular detection method based on dual-excitation ratio up-conversion fluorescent probes

Method used

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  • An Intracellular Detection Method Based on Dual Excitation Ratiometric Upconversion Fluorescent Probes
  • An Intracellular Detection Method Based on Dual Excitation Ratiometric Upconversion Fluorescent Probes
  • An Intracellular Detection Method Based on Dual Excitation Ratiometric Upconversion Fluorescent Probes

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0078] This example uses dye-sensitized Yb-doped up-conversion nanoparticles IR808-UCNP@Yb-F127 to detect hypochlorous acid:

[0079] (1) 1mmol oil-soluble nanoparticles NaYF 4 :Yb,Er@NaYF 4 : The dye IR808 of Yb and 1 μ mol is dispersed in 20mL solvent methylene chloride, mixes one hour, obtains solution one;

[0080] (2) Dissolve 400mg of F127 in 2mL of dichloromethane, ultrasonically mix for 2min, and add dropwise to solution 1 under constant stirring to obtain solution 2;

[0081] (3) The solution is rotated and evaporated to remove the solvent dichloromethane, and after the volatilization of the dichloromethane is completed, it is redispersed in the aqueous solution;

[0082] (4) After the step (3) is completed, centrifuge and wash twice with deionized water to obtain the water-soluble dye-sensitized up-conversion nanoprobe.

[0083] (5) reacting the nanoprobe in step (4) with the target detection substance (hypochlorous acid) of 0-100 μ M for 30 minutes;

[0084] (6)...

Embodiment 2

[0089] This example uses dye-sensitized Nd-doped up-conversion nanoparticles IR808-UCNP@Nd-F127 to detect hypochlorous acid:

[0090] (1) 1mmol oil-soluble nanoparticles NaYF 4 :Yb,Er@NaYF 4 : Nd and 1 μmol of the dye IR808 were dispersed in 20mL of dichloromethane, mixed for one hour to obtain solution 1;

[0091] (2) Dissolve 400mg of F127 in 2mL of dichloromethane, ultrasonically mix for 2min, and add dropwise to solution 1 under constant stirring to obtain solution 2;

[0092] (3) The solution is rotated and evaporated to remove the solvent dichloromethane, and after the volatilization of the dichloromethane is completed, it is redispersed in the aqueous solution;

[0093] (4) After step (3) is completed, centrifuge and wash twice with deionized water to obtain a water-soluble dye-sensitized up-conversion nanoprobe;

[0094] (5) reacting the nanoprobe in step (4) with the target detection substance (hypochlorous acid) of 0-100 μ M for 30 minutes;

[0095] (6) Measure t...

Embodiment 3

[0100] refer to image 3 , near-infrared dual-excitation confocal microscopy system, including 2 independent lasers (980nm laser 1 and 808nm laser 2 2), laser scanning confocal microscope and fluorescence signal acquisition device. Among them, the laser scanning confocal microscope includes basic components such as an oil mirror 5, a dichroic mirror 1 6, a dichroic mirror 2 7, and a mirror 8; the fluorescence signal acquisition device includes a spectrometer 3 and a CCD camera 4 (as an image controller) .

[0101] The excitation light emitted by the excitation light source is focused by the oil lens to the cell to be observed, and the up-conversion light emitted by the nano-probe in the cell to be tested passes through the oil lens 5, dichroic mirror 1 6, and dichroic mirror 2 7 in sequence. , mirror 8, lens one 11, pinhole 9, lens two 12, optical filter 10 and lens three 13, captured by spectrometer 3. The up-converted light emitted by the nano-probe in the cell to be teste...

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Abstract

The invention belongs to the technical field of nano biomaterials, and discloses an intracellular detection method based on a dual excitation ratio type up-conversion fluorescent probe. The method includes contacting and reacting a nanoprobe and a target to be measured in a cell, using double excitation light to detect the concentration of the target to be measured by monitoring the luminescence intensity of the nanoprobe; the nanoprobe is dye-sensitized rare earth doped on Converting nanoparticles; the target to be detected is selected from reactive oxygen species, reactive nitrogen species, or intracellular metal ions related to many physiology and pathology in cells. The method utilizes the energy transfer process of dyes to efficiently sensitize upconversion nanoparticles to improve the sensitivity of detection; a dual excitation ratio probe model is provided as a setting reference to reduce detection caused by complex intracellular environment, probe distribution, and equipment. Deviation; provide the design scheme of the near-infrared dual excitation confocal microscope system, and provide new ideas for the detection of intracellular substances.

Description

technical field [0001] The invention belongs to the technical field of nanobiological materials, and in particular relates to an intracellular detection method based on a double-excitation ratio type up-conversion fluorescent probe. Background technique [0002] The development of non-invasive fluorescent probes to monitor biomolecules or physiological processes in living cells is crucial to understanding cell biology, pathology, and other biomedical-related sciences. However, the current intracellular analysis is mainly through targeted imaging or non-quantitative fluorescent labeling, which cannot meet the actual needs. One reason for this is the low sensitivity of the probe. For example, the concentration of hypochlorous acid in cells is very low. If there is no external stimulus, it is difficult for general probes to detect endogenous hypochlorous acid. Although the in vitro detection limit of some dye probes can also reach very low, their poor water solubility and sho...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N21/64
CPCG01N21/6486
Inventor 卢珊陈学元柯建熙商晓颖刘䶮李幸俊宋晓荣
Owner FUJIAN INST OF RES ON THE STRUCTURE OF MATTER CHINESE ACAD OF SCI
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