Extracellular vesicle separation and enrichment method based on anionic polymer modified matrix

Abstract

The invention discloses a simple, low-cost and high-universality extracellular vesicle separation and enrichment method. The method is characterized in that extracellular vesicles in sample types suchas a culture medium, plasma, serum, urine, saliva, emulsion, pleural effusion and cerebrospinal fluid are separated and enriched through electrostatic interaction. According to the implementation mode of the method, the extracellular vesicles are captured by utilizing an anionic polymer modified matrix material under an acidic condition, and the extracellular vesicles captured in the previous step are eluted under a neutral or alkaline condition, so that the purpose of separating and enriching the extracellular vesicles is finally achieved. Compared with an existing extracellular vesicle separation and enrichment method, the method has the advantages of being simple in separation process, low in cost, good in universality and high in downstream detection compatibility.

Description

technical field [0001] The invention belongs to the field of in vitro diagnosis and targeted drug loading, and in particular relates to a method for separating and enriching extracellular vesicles. Background technique [0002] Extracellular vesicles are a kind of particles secreted by cells into the extracellular space and wrapped by phospholipid bimolecular membranes, with a diameter between 30nm and 2000nm. As an intercellular communication carrier, it is widely distributed in human body fluids such as blood, urine, pleural effusion, saliva, ascites, milk, nasal mucus and cerebrospinal fluid, etc. 2,3 . A large number of studies have proved that extracellular vesicles carry effector molecules such as various proteins, lipids, DNA, mRNA, microRNA, and non-coding RNA, and they communicate with neighboring cells and distant cells through these effector molecules. communication and material exchange 1,4-6 . In recent years, more and more studies have confirmed that extrac...

AI Technical Summary

Problems solved by technology

Ultracentrifugation is widely used in the separation of extracellular vesicles, but the separation process requires a large ultracentrifuge, and a large amount of impurity proteins and nucleic acids are inevitably co-separated, resulting in low purity of the isolated extracellular vesicles. The difference in vesicle size and density will easily lead to a series of problems such as the discarding of some extracellular vesicles with lower density and smaller particle size, and the inevitable damage of extracellular vesicles caused by excessive centrifugal force 13

Although the ultrafiltration method can separate extracellular vesicles with high purity, the requirements for equipment are relatively simple, but the capture capacity of the sample is small due to clogging, and it is easy to cause extracellular vesicles to be unavoidable due to shear force. damage, and easy adhesion to the filter membrane, resulting in reduced separation efficiency 13

The immunoaffinity capture method has the advantages of high separation efficiency and strong specificity, but has the disadvantages of high reagent cost, small capture capacity and poor versatility

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