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Rate-type rare-earth fluorescent probe and application in detecting bacillus-anthracis biomarker

A rare-earth fluorescence and fluorescent probe technology, applied in the field of analytical chemistry, can solve problems such as difficult to achieve accurate, simple, portable and rapid detection, complex pre-preparation, etc., to eliminate the influence of background fluorescence and scattered fluorescence, high sensitivity and selectivity , cheap effect

Inactive Publication Date: 2017-07-18
EAST CHINA NORMAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0005] The purpose of the present invention is to provide a rare earth metal ion-doped ratiometric fluorescent probe for the deficiencies in the prior art and the application of the probe for detecting DPA, so as to overcome the complicated early stage in the existing method for detecting DPA Preparing and using expensive large-scale instruments, it is difficult to achieve accurate, simple, portable and rapid detection of defects

Method used

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  • Rate-type rare-earth fluorescent probe and application in detecting bacillus-anthracis biomarker
  • Rate-type rare-earth fluorescent probe and application in detecting bacillus-anthracis biomarker
  • Rate-type rare-earth fluorescent probe and application in detecting bacillus-anthracis biomarker

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Embodiment 1

[0042] First prepare the fluorescent probe; add 500 μL of absolute ethanol containing 9.0 mg EDC and 2.8 mg NHS to 600 μL of 20 mM DPA solution, and stir for 40 minutes. Next, 100 μL of APTES was added and reacted for 100 minutes. Then, add 200 μL of 20 mM Tb(NO 3 ) 3 solution into the mixed solution. The resulting mixture was used as a precursor. Then, Tb / DPA@SiO was prepared by the inverse microemulsion method 2 . Use 1 mL of n-hexanol, 1 mL of Triton X-100 and 4 mL of cyclohexane to make a microemulsion and add it to 300 μL of the previously prepared Tb / DPA solution. After 40 minutes of continuous stirring, add 25 μL of ammonia to the solution aqueous solution (28%) and 85 μL of TEOS. The stirred reaction was continued for 24 hours. An equal volume of acetone was used to separate the nanoparticles from the microemulsion and centrifuged 3 times with ethanol and water washes. In order to prepare amino-modified Tb / DPA@SiO 2 -NH 2 , the Tb / DPA@SiO 2 Suspended in the ...

Embodiment 2

[0044] Mix 500 μL of 2 mM GMP solution and 1.0 mL of 0.5 mg / mL Tb / DPA@SiO 2 -NH 2 The solutions were mixed and then shaken for 30 minutes. Next, 500 µL of 2 mM Eu(NO 3 ) 3 Add to the mixed solution and shake at room temperature for 100 minutes. Then, the product was collected by centrifugation at 10000 rpm for 5 minutes, and washed 3 times with ultrapure water to remove unreacted reagents. Finally, the Tb / DPA@SiO 2 -Eu / GMP mixture stored in 2 mL H 2 O for further use. (The prepared material is characterized as figure 2 as shown, figure 2 (A) is Tb / DPA@SiO 2 -NH 2 The transmission scanning electron microscope characterization diagram of figure 2 (B) is Tb / DPA@SiO 2 -NH 2 The transmission scanning electron microscope characterization diagram of figure 2 (C) is Tb / DPA@SiO 2 -NH 2 , Tb / DPA@SiO 2 -Eu / GMP, Tb / DPA@SiO 2 - Infrared spectrum characterization of Eu / GMP / DPA, figure 2 (D) is Tb / DPA@SiO 2 - Fluorescence excitation EX and emission spectra EM) of Eu / ...

Embodiment 3

[0046] DPA was detected with the prepared fluorescent probe and the aforementioned detection conditions. Add 80 μL of Tris-HCl buffer (pH 7.4, 50 mM) to 10 μL of the prepared Tb / DPA@SiO 2 -Eu / GMP solution. Afterwards, 10 μL of DPA of different concentrations (0.25-20 µM) was added to the above solution. The final concentration of Tris-HCl was 40 mM, while the final concentration of DPA was in the range of 0.025-2.0 µM. In order to explore the selectivity of ratiometric fluorescent probes for detecting DPA, different materials were selected for interference experiments, including phenoxyacetic acid (POA), terephthalic acid (p-PA), benzoic acid (BA), cysteine amino acid (Cys), glutamic acid (Glu), the final concentration of these substances is 50 μM. Finally, the emission spectra were recorded under excitation at a wavelength of 272 nm. When the concentration is 0.025-2.0 µM, the fluorescence intensity is linearly related to the DPA concentration: Y=1.225X+0.033, R 2 =0.991...

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Abstract

The invention discloses a rate-type rare-earth fluorescent probe Tb / DPA@SiO2-Eu / GMP, a preparation method of visual test paper and an application thereof in the aspect of detecting bacillus-anthracis biomarker, namely 2, 6 dipicolinic acid (DPA). After the fluorescent probe is synthesized by a reverse microemulsion method, the quantitative detection for the concentration of the DPA is realized by utilizing a time-resolved fluorescence analysis method. Further, the portable visual test paper is prepared by soaking filter paper in a fluorescent probe solution, and can be applied to mobile-phone software for recognizing colors to detect the concentration of the DPA. The invention has the advantages that the currently-common traditional detection mode is changed, the complex operation process is not needed and the cost is saved. The invention has the characteristics of high accuracy, sensitivity and selectivity. The prepared test paper has the advantages of being convenient in carry and use, low in price and the like and has wide application prospect.

Description

technical field [0001] The invention relates to the technical field of analytical chemistry, in particular to a ratio-type fluorescent probe doped with double rare earth metal ions and the use of the probe in detecting the concentration of Bacillus anthracis biomarkers. Background technique [0002] Bacillus anthracis is a Gram-positive spore-forming aerobic bacterium. Anthrax is a major disease of herbivores. Exposure to spores in soil and on fur can cause infection. Spores can infect humans through respiratory tract, digestive tract and skin contact, and cutaneous anthrax is the most common. Due to the strong resistance of anthrax spores in the environment, it has been listed as the number one biological warfare agent in the military. As a main component of the shell of Bacillus anthracis, 2,6 dipicolinic acid (DPA) occupies the About 5-15% of the mass of dry Bacillus anthracis spores, so the content of DPA can be quantitatively detected to indirectly detect Bacillus anth...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N21/78G01N21/64
CPCG01N21/78G01N21/6486
Inventor 张闽王琦贤施国跃薛诗凡陈子晗
Owner EAST CHINA NORMAL UNIV
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