Method of selectively detecting thrombin by phosphorescent probe

A technology of thrombin and phosphorescence, applied in fluorescence/phosphorescence, biological testing, measuring devices, etc., to achieve the effects of long phosphorescence life, strong selectivity, and high phosphorescence intensity

Inactive Publication Date: 2018-08-07
TIANJIN NORMAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

RTP sensors mostly study pure quenching systems, and there are few studies on "off-on" systems

Method used

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  • Method of selectively detecting thrombin by phosphorescent probe
  • Method of selectively detecting thrombin by phosphorescent probe
  • Method of selectively detecting thrombin by phosphorescent probe

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0040] Synthesis of Mn-doped ZnS quantum dots coated with MPA

[0041] Add 5 mL of 0.1 M zinc sulfate, 5 mL of 0.01 M manganese acetate, and 50 mL of 0.04 M MPA into a 100 mL three-necked flask, and adjust the pH of the mixed solution to 11 with 1 M NaOH solution. The mixture was magnetically stirred at room temperature for 30 min in a nitrogen environment to ensure that the stabilizer MPA and Zn 2+ and Mn 2+ Fully complexed. Then add 5 mL of 0.1 M sodium sulfide aqueous solution with a syringe under the condition of air isolation, and continue stirring at room temperature for 20 min, then stop the N 2 . The three-neck flask was placed in the air, aged at 50 °C for 2 hours with constant stirring. MPA-coated Mn-doped ZnS phosphorescent quantum dots with room-temperature phosphorescent properties were obtained. Add absolute ethanol (3 times the amount of absolute ethanol), centrifuge at 12,000 rpm for 5 min at high speed, discard the supernatant, wash the obtained product r...

Embodiment 2

[0043] 1. Synthetic method with reference to embodiment 1;

[0044] 2. Using phosphorescence as the detection method, MPA-coated Mn-doped ZnS quantum dots are used for specific detection of thrombin:

[0045] ① Add 5 μL of MPA-coated Mn-doped ZnS quantum dot mother solution to the centrifuge tube in turn, and mix 50 μL (0.1M) pH=7.4 Tris-HCl buffer solution, then add 50 μL of 6 μM TBA solution as a phosphorescent quencher , add high-purity water to make up to 500 μL. After shaking well and standing still for 35 min, the fluorescence spectrophotometer was adjusted to the phosphorescence detection mode, and the excitation wavelength was set to 315 nm for detection.

[0046] ②Add 5 μL of MPA-coated Mn-doped ZnS quantum dot mother solution to the centrifuge tube in sequence, and mix 50 μL (0.1M) pH=7.4 Tris-HCl buffer solution, and then add 50 μL of 6 μM TBA solution as a phosphorescent quencher , add high-purity water to make up to 450 μL. Shake well and let it stand for 35 mi...

Embodiment 3

[0048] 1. Synthetic method with reference to embodiment 1;

[0049] 2. Using phosphorescence as the detection method, MPA-coated Mn-doped ZnS quantum dots are used for specific detection of thrombin:

[0050] ① Add 30 μL of MPA-coated Mn-doped ZnS quantum dot mother solution to the centrifuge tube, 50 μL (0.1M) pH=8.0 Tris-HCl buffer solution and mix well, then add 50 μL of 6 μM TBA solution as a phosphorescent quencher , add high-purity water to make up to 500 μL. After shaking well and standing still for 35 min, the fluorescence spectrophotometer was adjusted to the phosphorescence detection mode, and the excitation wavelength was set to 315 nm for detection.

[0051] ② Add 30 μL of MPA-coated Mn-doped ZnS quantum dot mother solution to the centrifuge tube, 50 μL (0.1M) pH=8.0 Tris-HCl buffer solution and mix well, then add 50 μL of 6 μM TBA solution as a phosphorescence quencher , add high-purity water to make up to 450 μL. Shake well and let it stand for 35 minutes. Aft...

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Abstract

The invention discloses a method of selectively detecting thrombin by a phosphorescent probe. The phosphorescent intensity of a system is recovered as Mn doped ZnS phosphorescent quantum dots coated by MPA and a thrombin adapter are mixed with surfaces, far from the quantum dots, of TBA in a quenching system because of relatively high specificity combination between a thrombin adapter (TBA) and thrombin. Related to the phosphorescent quantum dots, the phosphorescent quantum dots are applied more widely in detection as the phosphorescent service life is long, interference of autofluorescence and scattered light can be avoided, the selectivity is high, no deoxidant and inductor are needed to be added during detection and the like. The phosphorescent quantum dots and TBA are not modified, sothat the synthetic steps are simplified to a great extent, and meanwhile, the experimental cost is also lowered. The method is high in accuracy and high in sensitivity, and has a very good applicationprospect in detection of an actual sample.

Description

[0001] This patent is funded by the National Natural Science Foundation of China General Project 2137509, Tianjin Natural Science Foundation Youth Project (No.17JCQNJC05800) and Tianjin Normal University Doctoral Fund Project (No.52XB1510). technical field [0002] The invention belongs to the technical field of biological analysis and detection, and mainly relates to the application of a label-free phosphorescent probe to the quantitative detection of thrombin. Background technique [0003] In recent years, the fluorescence analysis method using doped quantum dots as markers has attracted extensive attention from researchers at home and abroad, and has become a hot spot in current scientific research. The introduction of impurity ions in quantum dots will change the optical, electrical and magnetic properties of quantum dots. The change of optical properties of quantum dots by impurity ions is particularly important because doping can induce some optical properties that are...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N21/64G01N33/573G01N33/86
CPCG01N21/64G01N33/573G01N33/86G01N2333/90G01N2800/224
Inventor 李妍熊艳叶雨萌张菲
Owner TIANJIN NORMAL UNIVERSITY
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