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Method for extracting human tumor cell membrane protein

A tumor cell and cell membrane technology, applied in the field of biological products, can solve the problems of incompatible enzymatic hydrolysis and mass spectrometry identification, poor solubilization of membrane proteins, and inability to effectively stabilize the biological activity of membrane proteins, etc., to achieve high surface activity and increase protein concentration , the effect of excellent physical and chemical properties

Inactive Publication Date: 2015-03-25
深圳市人口和计划生育科学研究所
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Problems solved by technology

However, due to their own characteristics, these surfactants have their own limitations in the specific application of membrane proteomics, such as poor solubilization of membrane proteins, inability to effectively stabilize the biological activity of membrane proteins, or incompatibility with subsequent enzymes. solution and mass spectrometric identification, etc.

Method used

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  • Method for extracting human tumor cell membrane protein
  • Method for extracting human tumor cell membrane protein
  • Method for extracting human tumor cell membrane protein

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Embodiment Construction

[0023] The present invention will be further described below in conjunction with accompanying drawings and examples.

[0024] A method for extracting human tumor cell membrane protein, comprising the following steps:

[0025] (1) Use a 10 mmol / L Tris-HCl solution on the collected human tumor cells, and use a Dounce homogenizer on ice to homogenize for 60 minutes;

[0026] (2) The homogenate was centrifuged at 4000 rpm for 15 minutes at 4°C, the supernatant was collected, and the precipitate was unbroken cells and cell nuclei and other organelles;

[0027] (3) The cell supernatant was centrifuged at 14400rpm at 4°C for 30 minutes, the supernatant was collected, and the cell organelles such as mitochondria were precipitated;

[0028] (4) The cell supernatant was centrifuged at 35300rpm at 4°C for 60 minutes, the precipitate was collected, and the supernatant was a solution such as cytoplasm;

[0029] (5) Membrane protein lysate (7mmol / L urea, 2mmol / L thiourea, 4% symmetrical s...

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Abstract

The invention discloses a method for extracting human tumor cell membrane protein. The method comprises steps of disrupting cells, carrying out differential centrifugation separation on a cell membrane, dissolving membrane protein and carrying out Western Blot detection on the purity of the membrane protein. Mainly aiming at the extraction of the human tumor cell membrane protein, the invention provides a new method for research on tumor cell membrane proteomics. The method disclosed by the invention is conducive to a further mechanism research on the influence of antitumor drugs on the expression level of the human tumor cell membrane protein, and a useful technical means is provided for the screening and related researches of antitumor drug targets.

Description

technical field [0001] The invention relates to the field of biological products, and specifically provides a method for extracting human tumor cell membrane proteins. technical background [0002] The cell membrane is the cell's barrier to the outside world and the interface between the internal and external environment of the cell. It isolates the cell from the surrounding environment and maintains a relatively stable intracellular environment. Membrane proteins are an important part of the cell membrane, performing important functions such as material exchange inside and outside the cell, cell recognition and immune response, signal transduction and regulation, and energy transfer. In the genome, genes encoding membrane proteins account for about one-third of the total number of genes. Among the known drug targets, about 70% are membrane proteins, including G protein-coupled receptors, ion channel proteins, transporters, cytochrome P450, etc. The dynamic observation and...

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Application Information

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IPC IPC(8): C07K14/47C07K1/34
CPCC07K14/47
Inventor 洪文旭欧阳建平邓晰月曾序春段山
Owner 深圳市人口和计划生育科学研究所
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