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Preparation method of temperature-sensitive response polymer array and application of temperature-sensitive response macromolecule array to pretreatment of samples

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 Eliminate sample loss and contamination, avoid loss, and improve detection sensitivity

Active Publication Date: 2017-09-12
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 response polymer array and application of temperature-sensitive response macromolecule array to pretreatment of samples
  • Preparation method of temperature-sensitive response polymer array and application of temperature-sensitive response macromolecule array to pretreatment of samples

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0025] Embodiment 1: in combination with figure 1 , detailing the preparation process of the controllable hydrophilic array of polymer PNIPAM.

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

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

Embodiment 2

[0031] Embodiment 2: in combination with figure 2 , taking the enzymatic hydrolysis solution of cytochrome C (cyt-c) as an example to illustrate the application of the PNIPAM controllable hydrophilicity array in protein / peptide self-desalting.

[0032] (1) Preparation of sample solution: 1mL containing 50mM NH 4 HCO 3 0.5mg / mL cyt-c solution with 100μL containing 50mM NH 4 HCO 3 0.2mg / ml trypsin solution was mixed and incubated in a 37° incubator for 24h. with 50mM NH 4 HCO 3The solutions were diluted so that their final concentrations were 4 pmol / μL, 400 fmol / μL, and 40 fmol / μL, respectively. (The sample in this embodiment is a salt solution of protein. To analyze each peptide after enzymolysis, the proteolytic must be enzymatically hydrolyzed into peptides with enzymes, and then desalted and enriched, mass spectrometry analysis; if the sample is pure protein Salt solution, then directly do not need to use enzyme treatment, after desalting, the mass spectrometry analy...

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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 method for preparing a temperature-sensitive responsive polymer array and its application in sample pretreatment, specifically comprising growing polymer poly(N-isopropylacrylamide) directly on the surface of an indium tin oxide glass slide (ITO glass slide) (PNIPAM) to prepare controllable hydrophilicity arrays and use them for sample pretreatment. Background technique [0002] Biomass spectrometry is an important technical means of proteomics research. During the processing of protein samples, the presence of non-volatile salts will inhibit the ionization of samples, resulting in a decrease in detection sensitivity or even failure to detect. Therefore, for protein or polypeptide samples Performing desalting is often an essential step prior to mass spectrometry analysis. [0003] At present, the desalting method for protein / peptide samples is mainly based on solid-phase extraction technology, that is, the protein or peptide...

Claims

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Application Information

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IPC IPC(8): G01N27/62G01N1/28C08F120/54
CPCG01N1/28G01N33/6851C08F120/54
Inventor 刘颖鞠熀先孟潇成全
Owner NANJING UNIV