Method and system for extracting hydrothorax exosome

A technology for exosomes and pleural effusion, applied in the field of rapid extraction of pleural effusion exosomes, can solve the problems of complex process and sample loss, and achieve the effect of complete structure, simple extraction method and high purity

Pending Publication Date: 2019-12-10
阔然生物医药科技(上海)有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0009] However, for other common clinical samples with more impurities and complex components such as pleural effusion, the extraction of exosomes by the above method usually requires an initial sample volume of more than tens of milliliters, or additional additions such as hydrophilic polymers are added during the extraction step. substances, or require multiple centrifugation and precipitation steps, which are complex and often result in large sample losses

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  • Method and system for extracting hydrothorax exosome
  • Method and system for extracting hydrothorax exosome
  • Method and system for extracting hydrothorax exosome

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0031] Refer to attached figure 1 , the method for extracting pleural effusion exosomes in the present application includes:

[0032] 5ml of frozen human pleural fluid stored at -80°C was thawed at room temperature;

[0033] After the pleural effusion is completely thawed, the pleural effusion is centrifuged to obtain the supernatant, and the centrifugal force is 16000g;

[0034] One end of the filter is sealed with the injection port of the syringe, the supernatant is transferred into the syringe, and then the plunger of the syringe is slowly pushed, and the syringe pushes the supernatant into the filter for filtration to obtain the filtrate, wherein the filter Capable of intercepting substances with a diameter of 0.45 μm;

[0035] Pass the filtrate through the QIAGEN exoEasy Maxi Kit exosome extraction kit, and extract exosomes according to the method on the product manual:

[0036] 1) Mix the filtrate with an equal volume of buffer XBP (provided by the kit);

[0037] 2)...

Embodiment 2

[0040] Refer to attached figure 1 , the method for extracting pleural effusion exosomes in the present application includes:

[0041] 10ml of frozen human pleural fluid stored at -80°C was thawed at room temperature;

[0042] After the pleural effusion is completely thawed, the pleural effusion is centrifuged to obtain the supernatant, and the centrifugal force is 12000g;

[0043] One end of the filter is sealed with the injection port of the syringe, the supernatant is transferred into the syringe, and then the plunger of the syringe is slowly pushed, and the syringe pushes the supernatant into the filter for filtration to obtain the filtrate, wherein the filter Capable of intercepting substances with a diameter of 0.45 μm;

[0044] Pass the filtrate through the QIAGEN exoEasy Maxi Kit exosome extraction kit, and extract exosomes according to the method on the product manual:

[0045] 1) Mix the filtrate with an equal volume of buffer XBP (provided by the kit);

[0046]2)...

Embodiment 3

[0049] Refer to attached figure 1 , the method for extracting pleural effusion exosomes in the present application includes:

[0050] 5ml of frozen human pleural fluid stored at -80°C was thawed at room temperature;

[0051] After the pleural effusion is completely thawed, the pleural effusion is centrifuged to obtain the supernatant, and the centrifugal force is 20000g;

[0052] One end of the filter is sealed with the injection port of the syringe, the supernatant is transferred into the syringe, and then the plunger of the syringe is slowly pushed, and the syringe pushes the supernatant into the filter for filtration to obtain the filtrate, wherein the filter Capable of intercepting substances with a diameter of 0.45 μm;

[0053] Pass the filtrate through the QIAGEN exoEasy Maxi Kit exosome extraction kit, and extract exosomes according to the method on the product manual:

[0054] 1) Mix the filtrate with an equal volume of buffer XBP (provided by the kit);

[0055] 2)...

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Abstract

The application provides a method and system for extracting hydrothorax exosome. The system comprises a centrifugal separator, filtration equipment and a centrifugal separation column, wherein the filtration equipment can intercept substances of which the diameter is 0.45 [mu] m; the centrifugal separator is used for performing centrifugal separation on hydrothorax to obtain supernatant; the filtration equipment is used for filtering the supernatant to obtain filtrate; and the exosome is extracted from the filtrate through a centrifugal column method. According to the system disclosed by the invention, by a method of performing centrifuging once and performing filtering once, a conventional membrane compatible centrifuging column reagent kit is combined, so that high-quality exosome can beeffectively extracted from hydrothorax as low as 5mL, the extracted exosome is complete in structure and high in purity, and can be directly used for downstream detection experimental researches including physical and chemical detection, exosome nucleic acid extraction and the like, and therefore, the method for quickly, easily and conveniently extracting the hydrothorax exosome is provided.

Description

technical field [0001] The invention relates to a method for extracting biological materials, in particular to a new method and system for rapidly extracting pleural effusion exosomes. Background technique [0002] Exosomes are a multivesicular body with a diameter of about 40-100nm, which is secreted by the cells themselves or released when the cells die. Many tissues such as blood, saliva, urine, milk, pleural effusion (pleural effusion), ascites Natural exosomes exist in the interstitial fluid. Exosomes contain various molecules such as proteins, lipids, DNA, mRNA, and miRNA from the original cells. These biomolecules participate in the occurrence, growth, invasion, and metastasis of tumors and other diseases through the circulatory system. Diagnostic biomarkers are of great significance in clinical diagnosis and treatment. For example, Wang Yan reported the study of exosomal miRNA as three kinds of markers of pleural effusion after bathing (Nanchang University Master R...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/071
CPCC12N5/0602C12N2509/10
Inventor 陈鑫泽
Owner 阔然生物医药科技(上海)有限公司
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