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Preparation and application of type I hyaluronidase fluorescent nano sensor

A hyaluronidase and nanosensor technology, applied in the fields of nanotechnology and analysis and detection, to achieve the effects of mild conditions, high specificity, good social value and application prospects

Active Publication Date: 2021-01-05
CHANGSHA UNIVERSITY OF SCIENCE AND TECHNOLOGY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, this sensing system has not been reported in the detection of Hyal-1 in urine

Method used

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  • Preparation and application of type I hyaluronidase fluorescent nano sensor
  • Preparation and application of type I hyaluronidase fluorescent nano sensor
  • Preparation and application of type I hyaluronidase fluorescent nano sensor

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0031]Example 1: Preparation and characterization of nanosensors

[0032]Add 1 mg of TPC3 dye and 2 mg of CHA polymer to 1 ml of DMSO, slowly drop 3 ml of ultrapure water into the solution under magnetic stirring at 37°C, react for 30 minutes, and white micelle particles gradually appear in the solution. After the reaction, the reaction solution was used to remove the unwrapped TPC3 dye molecules with a molecular weight cut-off 3500KD dialysis bag, and dialyzed for 3 days. During this process, the white micellar particles were slowly dispersed in the dialysis bag until they disappeared to prepare a TPC3@CHA micellar solution of uniform size. The final product was stored at 4°C. byfigure 1 (A) The transmission electron microscope image shows that the size of TPC3@CHA is about 160nm, and there is no agglomeration phenomenon, and the uniformity is good.

Embodiment 2

[0033]Example 2: Titration curve of nano sensor response

[0034]The titration curve is determined under optimized conditions.figure 2 (A) shows that after different concentrations of Hyal-1 (0-12μg / mL), the fluorescence intensity of the nanosensor increases significantly with the increase of Hyal-1 concentration. And fromfigure 1 (B) It can be seen that after the nanosensor responds to Hyal-1, the size of the nanosensor after being degraded by Hyal-1 is about 5nm.figure 2 (B) shows that the signal-to-background ratio (S / B) of the nanosensor and the analyte Hyal-1 in the range of 0.25-6.0μg / mL shows a good linear relationship, and the linear function is y = 1.267x + 2.1811;R2 = 0.9956. The probe can accurately detect Hyal-1 in urine.

Embodiment 3

[0035]Example 3: Selectivity investigation of nano sensor

[0036]Take the nanosensor and add it to the PB buffer at a concentration of 0.02 mmol / L TPC3@0.3mg / mL CHA, and then add various representative interference species (0.1 μM / mL H2O2, Urea, Uric acid, Glu, Gly, Cys,Glucose, VC, GSH, Cathepsin, Trypsin, Thrombin, Galactosidase, Ribnudease, Hyal-2 (5μg / mL), Hyal-3 (5μg / mL ), Hyal-4( 5μg / mL), PH-20 (5μg / mL) and Hyal-1(5μg / mL)). At the same time, add a blank group, add only 2 μL PBS buffer solution, shake well, measure and record the fluorescence intensity of each group of solutions at excitation wavelength = 375 nm and emission wavelength = 495 nm, to obtain various species of nano-sensing Influence of fluorescence signal. The signal-to-background ratio is the fluorescence value of the solution after adding various interfering species divided by the fluorescence value of the blank group, the experiment is repeated three times, and the average value is taken.

[0037]Result analysis: su...

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Abstract

The invention discloses a polymer nano sensor and application of the polymer nano sensor in detection of hyaluronidase type 1 (Hyal-1). Based on the principle that different hyaluronidase differentially degrades hyaluronic polymer nano-micelles, hydrogen bond induced enhanced dye molecules are wrapped in nano-micelle particles, and a high-selectivity and high-sensitivity Hyal-1 fluorescent nano-sensor is designed by utilizing stimulus-responsive nano-materials to adjust the interaction between dye and water molecules. A type I hyaluronidase detection kit comprises a type I hyaluronidase fluorescent nano sensor and a solvent. Results show that the kit has the advantages of high specificity, good sensitivity, simplicity and convenience in operation, economy, practicability and the like; andthe detection of Hyal-1 in an actual urine sample is realized. Therefore, the method provided by the invention has original innovativeness, good social values and application prospects, and is expected to be applied to detection and analysis of Hyal-1 in urine of bladder cancer patients.

Description

Technical field[0001]The invention belongs to the field of nanotechnology and analysis and detection, and relates to the construction of a type 1 hyaluronidase fluorescence nano sensor and a type 1 hyaluronidase fluorescence nano sensor platform based on the principle of hydrogen bond induction and enhancement.Background technique[0002]Hyaluronidases (HAases) belonging to the endoglycosidase family can degrade hyaluronic acid (HA) into oligosaccharide fragments to maintain various physiological functions such as angiogenesis, fertilization, embryonic development and cell movement. HAases have subtypes with similar gene sequences but different functions. One type of hyaluronidase (Hyal-1) is usually present in the lysosome to catalyze the degradation of intracellular HA to maintain key physiological functions. When the body's function is abnormal or diseased, Hyal-1 in the cell will be overexpressed and spilled into the tissue and body fluid. Hyal-1 in body fluids has become one of t...

Claims

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Application Information

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IPC IPC(8): G01N21/64B82Y15/00
CPCG01N21/6428B82Y15/00
Inventor 杨盛罗金秋杨荣华
Owner CHANGSHA UNIVERSITY OF SCIENCE AND TECHNOLOGY
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