Preparation method of saliva extracellular vesicles and application of saliva extracellular vesicles to molecular diagnosis

A salivary exosome and saliva technology, applied in the biological field, can solve problems such as background impurities and interference that cannot be solved, and achieve the effects of improving extraction efficiency, removing interference and high recovery rate.

Active Publication Date: 2016-06-15
SHANGHAI JIAO TONG UNIV
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Problems solved by technology

Although the precipitation method improves the extraction efficienc

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  • Preparation method of saliva extracellular vesicles and application of saliva extracellular vesicles to molecular diagnosis
  • Preparation method of saliva extracellular vesicles and application of saliva extracellular vesicles to molecular diagnosis
  • Preparation method of saliva extracellular vesicles and application of saliva extracellular vesicles to molecular diagnosis

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Embodiment

[0033] 1. Extraction of salivary exosomes

[0034] The preparation method of salivary exosomes is as follows: figure 1 As shown, the saliva samples of adult healthy volunteers and lung cancer patients were collected, and background interfering proteins such as high-abundance proteins and mucins were removed by affinity chromatography and membrane separation technology. The specific method was to use 1:1 starch and Prepare the stationary phase of the affinity chromatography column according to the protein ratio, and then connect the 5 μm PVDF membrane to complete the integration of chromatography and membrane separation technology. Firstly, the mobile phase PBS was used to equilibrate the column and activate the membrane. Then, a corresponding proportion of saliva samples was injected into the injection port of the chromatogram, and the saliva samples were eluted with a mobile phase PBS with a total volume of 2 mL at a speed of 800 μL / min. The eluted samples were collected an...

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Abstract

The invention discloses a preparation method of saliva extracellular vesicles and application of the saliva extracellular vesicles to molecular diagnosis.The preparation method includes the steps of removing high-abundance proteins from saliva samples, filtering out mucoproteins, separating and extracting the extracellular vesicles, characterizing the obtained extracellular vesicles, extracting extracellular vesicle proteins, identifying the extracellular vesicle proteins and applying the extracellular vesicle proteins to lung cancer biomarker discovery.Currently, a normalized saliva extracellular vesicle preparation technology is absent, and high-abundance protein interference and background impurity interference, existing in existing preparation technologies, result in shortcomings of low extracellular vesicle recovery rate, fewer identified proteins and the like, so that true extracellular vesicle proteomics information can be reflected difficultly.The preparation method of the saliva extracellular vesicles and application of the saliva extracellular vesicles to molecular diagnosis have the advantages that saliva of healthy adult volunteers and lung cancer patients is taken as the samples, and an affinity chromatography and membrane separation combined technology is adopted, so that after the high-abundance proteins and the mucoproteins of the saliva are removed, the extracellular vesicles in the saliva are extracted centrifugally, proteomes of the extracellular vesicles are analyzed and more extracellular vesicle proteins are identified.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a preparation method of salivary exosomes and its application in molecular diagnosis. Background technique [0002] Exosomes (Extracellular Vesicles) refer to a vesicle with a membrane structure secreted by cells into the extracellular microenvironment. They are tiny particles containing membrane components released by cells during outward budding and cell membrane fission. Carry a variety of specific proteins, microRNA, lipids and other substances. The size of exosomes is between 0.05 and 1.00 μm. Cancer mechanism studies in recent years have confirmed that exosomes are related to the transmission of material, energy, and information between cells, especially between cancer cells or between cancer cells and normal cells. affect each other. Exosomes widely exist in tissues and body fluids, and play an important guiding role in the clinical diagnosis of diseases, predict...

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Application Information

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IPC IPC(8): G01N30/14
CPCG01N30/14G01N2030/146
Inventor 肖华孙妍夏志军尚志孙凯博曹成喜樊柳荫
Owner SHANGHAI JIAO TONG UNIV
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