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Protein-inorganic nanoflower with controllable protein distribution and size as well as preparation method and application of protein-inorganic nanoflower

A protein distribution and inorganic nanotechnology, applied in the field of biosensors, can solve problems such as poor repeatability of analysis results, achieve effective immobilization, good biocompatibility, and improve sensitivity

Pending Publication Date: 2022-05-27
JILIN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0005] The PIHN currently studied is mainly prepared by the method of blending proteins or biomacromolecules, and the reproducibility of the analysis results is poor.

Method used

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  • Protein-inorganic nanoflower with controllable protein distribution and size as well as preparation method and application of protein-inorganic nanoflower
  • Protein-inorganic nanoflower with controllable protein distribution and size as well as preparation method and application of protein-inorganic nanoflower
  • Protein-inorganic nanoflower with controllable protein distribution and size as well as preparation method and application of protein-inorganic nanoflower

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0029] Embodiment 1: A preparation method of protein-inorganic nanoflowers with controllable protein distribution and size, comprising the following steps:

[0030] (1) FITC-labeled HRP (FITC-HRP) and RhB-labeled Ab 2 (RhB-Ab 2 ) preparation method refer to the literature Chem.Mater.2018, 30, 1069-1077. Next, use FITC-HRP and RhB-Ab 2 Protein-inorganic nanoflowers were synthesized. First, 30 μL, 200 mmoL -1 The copper sulfate aqueous solution was added to the centrifuge tube, to which was added 1 mL containing 1.0 mg mL - 1 Phosphate buffer solution of FITC-HRP (0.01 mmol -1 , pH 7.4). After vortexing for 5 minutes, it was left to react at 4°C for 12 hours.

[0031] (2) After taking out, at 1000r min -1 Centrifuge for 1 min, dilute to 1 mL with deionized water, and set it at 1000 rpm. -1 Centrifuge again for 1 min at rpm and remove the supernatant. Then, 1 mL containing 0.25 mg mL was added thereto -1 RhB-Ab 2 phosphate buffered solution (0.01 mmol -1 , pH 7.0). ...

Embodiment 2

[0032] Embodiment 2: A preparation method of protein-inorganic nanoflowers with controllable protein distribution and size, comprising the following steps:

[0033] (1) First, 30 μL, 200 mmol -1 The copper sulfate aqueous solution was added to the centrifuge tube, and 1 mL containing 0.1 mg mL was added to it. -1 Phosphate buffer solution of HRP (0.01 mmol -1 , pH 7.4). After vortexing for 5 minutes, it was left to react at 4°C for 12 hours.

[0034] (2) After taking out, at 1000r min -1 Centrifuge for 1 min, dilute to 1 mL with deionized water, and set it at 1000 rpm. -1 Centrifuge again for 1 min at rpm and remove the supernatant. Then, 1 mL containing 0.25 mg mL was added thereto -1 Ab 2 phosphate buffered solution (0.01 mmol -1 , pH 7.0). After 5 minutes of vortexing, it was left to react at 4°C for 24 hours. Then at 1000r min -1 Centrifuge twice at a rotating speed for 2 minutes each time; then dilute to 1 mL with deionized water to obtain a protein-inorganic n...

Embodiment 3

[0035] Embodiment 3: A preparation method of protein-inorganic nanoflowers with controllable protein distribution and size, comprising the following steps:

[0036] (1) First, 30 μL, 200 mmol -1 The copper sulfate aqueous solution was added to the centrifuge tube, to which was added 1 mL containing 1.0 mg mL -1 Phosphate buffer solution of HRP (0.01 mmol -1 , pH 7.4). After vortexing for 5 minutes, it was left to react at 4°C for 12 hours.

[0037] (2) After taking out, at 1000r min -1 Centrifuge for 1 min, dilute to 1 mL with deionized water, and set it at 1000 rpm. -1 Centrifuge again for 1 min at rpm and remove the supernatant. Then, 1 mL containing 0.25 mg mL was added thereto -1 Ab 2 phosphate buffered solution (0.01 mmol -1 , pH 7.0). After 5 minutes of vortexing, it was left to react at 4°C for 24 hours. Then at 1000r min -1 Centrifuge twice at a rotating speed for 2 minutes each time; then dilute to 1 mL with deionized water to obtain a protein-inorganic nan...

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Abstract

The invention discloses a protein-inorganic nanoflower with controllable protein distribution and size, a preparation method of the protein-inorganic nanoflower and application of the protein-inorganic nanoflower as a detection reagent in imidacloprid immunoassay, and belongs to the technical field of biosensors. The protein-inorganic hybrid nanoflower is synthesized by coprecipitation of a copper sulfate solution, horse radish peroxidase, goat anti-mouse IgG and a phosphate buffer solution, so that the stability of a biological enzyme and an antibody is effectively enhanced, and the protein-inorganic hybrid nanoflower has the characteristics of biological signal specific recognition and biological signal amplification. According to the invention, the protein distribution and size regulation of the protein-inorganic hybrid nanoflower are realized. The immunoassay of imidacloprid is realized by using a competitive immunoassay method, and the sensitivity of pesticide analysis is greatly improved. The invention provides a method for preparing a nano structure which is simple in preparation, good in biocompatibility, large in specific surface area and tunable, high-flux, high-selectivity and high-sensitivity analysis of imidacloprid is realized, and a new view angle is provided for monitoring food, environment and public safety.

Description

technical field [0001] The invention belongs to the technical field of biosensors, and in particular relates to a protein-inorganic nanoflower with controllable protein distribution and size, a preparation method and its application as a detection reagent in imidacloprid immunoassay. Background technique [0002] The current levels of many disease markers are far below the monitoring capabilities of existing methods, so the establishment of ultrasensitive bioanalytical techniques is particularly important for safeguarding human health. The ELISA method has good selectivity and specificity, and is considered to be the most popular bioanalytical method in the fields of clinical diagnosis and food quality control. The method is based on the specific recognition principle between antigen and antibody, and its performance is related to the activity and content of biological enzymes. However, biological enzymes are easily affected by the external environment, which greatly limits...

Claims

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Application Information

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IPC IPC(8): G01N33/535G01N33/543G01N33/53G01N33/58G01N21/31G01N21/78
CPCG01N33/535G01N33/54306G01N33/5308G01N33/581G01N21/31G01N21/78Y02A50/30
Inventor 卢革宇闫旭苏丹丹孙鹏刘方猛王晨光刘晓敏贾晓腾
Owner JILIN UNIV
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