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Preparation method of temperature-sensitive responsive polymer array and its application in sample pretreatment

A polymer and array technology, applied in the preparation of temperature-sensitive responsive polymer arrays and its application in sample pretreatment, can solve the problems of sample enrichment and detection sensitivity limitations, lack of surface array convergence effect, etc., to achieve Avoid loss, fast detection, simple effect of preparation

Active Publication Date: 2019-07-23
NANJING UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In recent years, PNIPAM-modified nanoparticles have been used for the enrichment and release of temperature-controlled samples, but there are still limitations in the enrichment and detection sensitivity of samples due to the lack of converging effects of surface arrays.

Method used

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  • Preparation method of temperature-sensitive responsive polymer array and its application in sample pretreatment
  • Preparation method of temperature-sensitive responsive polymer array and its application in sample pretreatment
  • Preparation method of temperature-sensitive responsive polymer array and its application in sample pretreatment

Examples

Experimental program
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Effect test

Embodiment 1

[0025] Example 1: Attachment figure 1 , Detail the preparation process of the controllable hydrophilic array of polymer PNIPAM.

[0026] (1) The ITO glass slide was sprayed alternately with polyacrylamine hydrochloride (PAH) and sodium silicate solution, rinsed with ultrapure water, dried with nitrogen, and calcined in a resistance furnace at 450° for 4 hours. 100μL silanization reagent (O-(propargyl)-N-(triethoxysilylpropyl)carbamate) and 0.5g MgSO 4 ·7H 2 O modifies the alkyne group on the surface of the ITO glass slide through the vapor deposition reaction.

[0027] (2) Drop 50vol% 1-thioglycerol and 2wt% benzoin diethyl ether (DMPAP) in tetrahydrofuran solution on the above ITO glass slide, spread a photomask at 365nm 14.4mW·cm -2 Irradiate under ultraviolet light for 20s, rinse with tetrahydrofuran, ultrapure water, and blow dry with nitrogen (that is, 1-thioglycerol passes through the photoinitiator benzoin diethyl ether to make the sulfhydryl group covalently bond with the al...

Embodiment 2

[0031] Example 2: Attachment figure 2 Take the enzymatic hydrolysis solution of cytochrome C (cyt-c) as an example to illustrate the application of the controllable hydrophilic array of PNIPAM in the self-desalting of proteins / peptides.

[0032] (1) Prepare sample solution: put 1mL containing 50mM NH 4 HCO 3 0.5mg / mL cyt-c solution and 100μL containing 50mMNH 4 HCO 3 The 0.2mg / ml trypsin solution was mixed and incubated in a 37° incubator for 24 hours. Use 50mM NH 4 HCO 3 The solutions were diluted so that their final concentrations were 4pmol / μL, 400fmol / μL, and 40fmol / μL, respectively. (The sample in this example is a protein salt solution. To analyze each peptide after enzymatic hydrolysis, the protease must be digested into peptides with an enzyme, and then desalting and enrichment, and mass spectrometry analysis; if the sample is pure protein Salt solution, then directly do not use enzyme treatment, after desalting, mass spectrometry analysis is the mass-to-charge ratio of...

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Abstract

The invention relates to a preparation method of a temperature-sensitive response polymer array and application of the temperature-sensitive response macromolecule array to pretreatment of samples. Based on a shadow printing technology and an alkynyl and sulfydryl click chemistry technology, a hydroxyl hydrophilic microarray is established on an ITO slide, and then a controllable hydrophilic array of polymer poly-(N-isopropyl acrylamide) (PNIPAM) is generated by surface-initiated atom transfer radical polymerization in the hydrophilic microarray. When the temperature is lower than the critical temperature, due to the hydrophilic interaction of the polymer array, the samples are gathered on the surface of slide; when the temperature is higher than the clinical temperature, a protein / peptide fragment is absorbed by a hydrophobic polymer array due to the hydrophobic interaction, and salt and pollutants coexisting are washed away, so that the pretreatment of on-target self-desalting and efficient enrichment is performed on the samples, and MALDI-MS one-step rapid detection is performed on the target samples. The preparation method is simple in preparation and operation, high in universality, short in analysis time and low in cost, therefore, the preparation method has a certain biomedical application value.

Description

Technical field [0001] The invention relates to a preparation method of a temperature-sensitive polymer array and its application in sample pretreatment, specifically including directly growing polymer poly(N-isopropylacrylamide) on the surface of indium tin oxide glass (ITO glass) (PNIPAM) to prepare a controllable hydrophilic array and use it for sample pretreatment. Background technique [0002] Biological mass spectrometry is an important technical means for proteomics research. The presence of non-volatile salts in the processing of protein samples will inhibit the ionization of samples, resulting in a decrease in detection sensitivity or even failure to detect. Therefore, for protein or peptide samples Desalting is often an essential step before mass spectrometry analysis. [0003] At present, the method of desalting protein / peptide samples is mainly based on solid phase extraction technology, that is, the protein or peptide is retained on the adsorption material, and the sa...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N27/62G01N1/28C08F120/54
CPCC08F120/54G01N1/28G01N33/6851
Inventor 刘颖鞠熀先孟潇成全
Owner NANJING UNIV