Method for rapidly extracting exosome in plasma and extraction reagent

Abstract

The invention discloses a method for rapidly extracting exosomes in plasma, the extraction method comprises the following steps: S1, dropwise adding a reagent, S2, carrying out preliminary centrifugal precipitation, S3, carrying out secondary centrifugal precipitation, and S4, re-suspending the exosomes, specifically, S1, dropwise adding the reagent: adding an extraction reagent into a plasma sample, shaking and uniformly mixing, and standing for 0.5-5 minutes in a sterile environment at the temperature of 2-8 DEG C to obtain a mixed solution A; after the PEG reagent is added into the plasma and uniformly mixed, the plasma only needs to stand for 2 minutes at the temperature of 2-8 DEG C, so that the operation time is greatly shortened, the extraction speed is increased, the requirement on timeliness in clinical detection application is met, after standing, only 1000g centrifugation is needed for 5 minutes, the centrifugation time is short, the centrifugal force is small, the experimental time is further shortened, and the experimental efficiency is improved. The exosome precipitate is easy to dissolve due to small centrifugal force, and the relatively complete exosome is easy to obtain.

Description

technical field [0001] The invention relates to the technical field of biological separation and extraction, in particular to a method for rapidly extracting exosomes in plasma and an extraction reagent. Background technique [0002] Exosomes refer to small membrane vesicles (30-150nm) containing complex RNAs and proteins. Various cells can secrete exosomes under normal and pathological conditions. They are mainly derived from intracellular lysosomal particles formed by invagination. Multivesicular bodies, which are released into the extracellular matrix after fusion of the outer membrane of the multivesicles with the cell membrane, are naturally present in body fluids, including blood, saliva, urine, cerebrospinal fluid and milk, and exosomes are considered as Specifically secreted membrane vesicles that are involved in intercellular communication, there is growing interest in exosomes, whether to study their function or understand how to use them in the development of mini...

AI Technical Summary

Problems solved by technology

[0004] In the prior art, methods such as centrifugation, immune capture, or filtration are generally used to separate and purify exosomes in blood or body fluids. The centrifugal separation method will cause a certain degree of mechanical damage to the membrane structure of exosomes, which will affect subsequent analysis and research. And the cumbersome centrifugation operation limits the throughput of liquid biopsy. The separation method of immune capture requires the use of antibodies, which greatly increases the cost of sample processing, and the elution conditions after immune capture may affect the activity of exosomes. The filtration separation method can effectively separate components of different sizes in body fluids. However, some components in biological samples are easy to be enriched on the filter membrane, and the components larger than the membrane pore size will block the membrane pores, and the clogging of the membrane pores will make the pores smaller than Components of membrane pore size cannot effectively pass through the filter membrane, which will also cause excessive local pressure and even cause the filter membrane to rupture

Due to the long time and high centrifugation speed of exosomes extracted by precipitation method, there is a problem that exosomes are not easy to dissolve

Images