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Exosome separation method and separation device thereof

A separation method and exosome technology, which can be used in sterilization methods, methods of supporting/immobilizing microorganisms, semi-permeable membrane separation, etc., which can solve the problems of uneven particle size, low purity and recovery rate, and many impurity proteins. , to achieve the effect of reducing the possibility, high through-hole ratio, and high efficiency

Inactive Publication Date: 2018-11-23
SHENZHEN TOPMEMBRANES INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

There are many problems in the use of PEG to precipitate exosomes: such as low purity and recovery, more impurities (false positives), uneven particle size, difficult to remove polymers, mechanical force or chemical additives such as Tween-20 Will destroy exosomes, etc., so the effect is easily questionable
However, the purification effects of various products on the market vary, and there is still no absolute method or kit that can meet all requirements and isolate ideal exosomes from various samples

Method used

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  • Exosome separation method and separation device thereof
  • Exosome separation method and separation device thereof
  • Exosome separation method and separation device thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0039] The steps of the method for separating exosomes in serum are as follows:

[0040] (1) Serum is prepared by filtration or centrifugation;

[0041] (2) Remove platelets and cell debris contained in the serum by microfiltration membrane filtration with a pore size of 1 μm, and collect the filtrate;

[0042] (3) adopting the filtrate collected in the uniform aperture anodic aluminum oxide membrane filtration step (2) with an aperture of 300nm through oxygen plasma cleaning in advance for 3 minutes;

[0043] (4) Using the exosome collection surface to collect the filtrate obtained in step (3) to obtain exosomes with a diameter below 300 nm.

[0044] Such as Figure 5 Shown is a schematic diagram of the process of isolating exosomes, that is, the serum 5 containing platelets and cell debris is first filtered through a microfiltration membrane 6 to obtain a filtrate 7, which is then filtered through an anodized aluminum oxide membrane 8 with a uniform pore size, and exosomes...

Embodiment 2

[0046] The steps of the method for separating exosomes in serum are as follows:

[0047] (1) Serum is prepared by filtration or centrifugation;

[0048] (2) The serum in the step (1) is filtered by an anodic aluminum oxide membrane with a uniform aperture of 300 nm and a thickness of 60 μm soaked in hydrogen peroxide solution for 20 hours in advance, and the filtrate is collected;

[0049] (3) adopting the filtrate collected in the uniform aperture anodic aluminum oxide membrane filtration step (2) that is 30nm and thickness is 50 μm through the aperture of soaking in hydrogen peroxide solution for 15 hours in advance;

[0050] (4) Collect the exosomes on the anodic aluminum oxide film with a uniform pore size of 30 nm in step (3) to obtain exosomes with a diameter of 30-300 nm.

[0051] Such as Figure 6 Shown is the particle distribution diagram of exosomes isolated from serum by the method of this example, and it can be seen that the diameter of exosomes is concentrated b...

Embodiment 3

[0053] The steps of the method for separating exosomes in urine are as follows:

[0054] (1) Collect urine filtrate or supernatant by filtration or centrifugation;

[0055] (2) The filtrate or supernatant in step (1) is filtered by anodic aluminum oxide film with uniform aperture of 300nm and thickness of 30 μm through oxygen plasma cleaning for 5 minutes in advance, and the filtrate is collected;

[0056] (3) adopting the filtrate collected in the uniform aperture anodic aluminum oxide membrane filtration step (2) that is 30nm and thickness is 10 μm through oxygen plasma cleaning in advance for 4 minutes;

[0057] (4) Collect the exosomes on the anodic aluminum oxide film with a uniform pore size of 30 nm in step (3) to obtain exosomes with a diameter of 30-300 nm.

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Abstract

The invention provides an exosome separation method and a separation device thereof. The exosome separation method comprises the steps: firstly filtering or centrifuging blood, urea, saliva, cerebrospinal fluid, milk, ascetic fluid or amniotic fluid to remove impurities larger than 1 micrometer; then utilizing an even pore diameter anodized aluminum oxide film with the pore diameter correspondingto the diameter size of the exosome to filter, collecting filtrate or exosome on the even pore diameter anodized aluminum oxide film to obtain required exosome. The exosome separation device comprisesa membrane filter with the even pore diameter anodized aluminum oxide film as a filter membrane. By means of the separation method and the separation device thereof disclosed by the invention, the exosome can be simply and efficiently separated, the different sizes of exosome can be cut out and classified, and the different sizes of exosome can be respectively and correspondingly analyzed, researched and applied.

Description

technical field [0001] The invention relates to a method for separating exosomes and a separation device thereof. Background technique [0002] Discovered in 1986, exosomes are double-membrane vesicles with a diameter of about 30-300 nm, which can be produced by various cells in the body such as immune cells, stem cells, cardiovascular cells, reticulocytes, Platelets, nerve cells and tumor cells are actively secreted and widely distributed in peripheral blood, urine, saliva, cerebrospinal fluid, breast milk, ascites, amniotic fluid and other body fluids. [0003] Exosomes carry a large number of specific proteins (such as cytokines, growth factors) and functional mRNAs, miRNAs and other biologically active substances, and participate in physiological processes such as cell communication, cell migration, angiogenesis and anti-tumor immunity in the body. The occurrence and process of many diseases are closely related. Due to the special structure and function of exosomes, it...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/071C12M1/12B01D71/02B01D67/00
CPCB01D67/0039B01D71/022C12M47/04C12N5/0602
Inventor 郭秋泉赵呈春杨军
Owner SHENZHEN TOPMEMBRANES INC
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