A magnetic bead-based method for isolating ecDNA from exosomes

A technology for magnetic bead separation and exosomes, which is applied in the field of molecular biology, can solve the problems of complex experiments, time-consuming, and inability to perform automated operations, and achieve the effect of saving experimental manpower and simple operation methods

Active Publication Date: 2022-04-15
SHANGHAI CHANGZHENG HOSPITAL +1
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  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0006] However, there are many disadvantages in using cesium chloride gradient centrifugation to extract ecDNA. For example, cesium chloride and ethidium bromide are toxic reagents used in the extraction experiment, which may cause harm to the operator's body and the environment; Many, but also need to use ultracentrifuge, and ecDNA layered absorption technology is difficult, the overall experiment is complicated, and must be operated by trained professionals; the experiment is time-consuming and costly, and cannot be automated

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  • A magnetic bead-based method for isolating ecDNA from exosomes
  • A magnetic bead-based method for isolating ecDNA from exosomes
  • A magnetic bead-based method for isolating ecDNA from exosomes

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Embodiment Construction

[0054] In order to understand the content of the present invention more clearly, it will be described in detail with reference to the drawings and embodiments.

[0055] Such as figure 1 Shown is a schematic flow chart of the method for separating ecDNA in exosomes based on magnetic beads in the present invention, which specifically includes the following steps:

[0056] Exosomes are isolated from biological samples using conventional methods (such as magnetic bead method);

[0057] Using a lysate to lyse the isolated exosomes;

[0058] Use the magnetic bead suspension II to perform nucleic acid adsorption on the products after exosome lysis;

[0059] Washing the magnetic beads adsorbed with nucleic acid using washing solution II with a pH of 7 to 12;

[0060] Using washing solution III containing protease to wash the magnetic beads that have been washed with washing solution II;

[0061] The eluent is used to elute the magnetic beads that have been washed with the washing ...

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Abstract

The present invention relates to a method for separating ecDNA in exosomes based on magnetic beads. The ecDNA in exosomes can be separated with high selectivity through the magnetic bead method, without using complex operations such as ultracentrifugation, and avoiding the use of toxic reagents such as cesium chloride. It can adapt to automatic operation to separate and obtain high-purity ecDNA products, which greatly improves the experimental efficiency and effectively saves manpower and time costs.

Description

technical field [0001] The invention relates to the field of molecular biology technology, in particular to a method for isolating ecDNA in exosomes based on magnetic beads. Background technique [0002] Exosome (exosome) is a 30-150nm size, extremely low-density extracellular nanoscale capsule structure, which is formed by cells through a series of regulatory processes such as "endocytosis-fusion-efflux". Exosomes did not attract the attention of researchers for a long time after they were discovered. Until the introduction of nano-analysis technology and the announcement of the Nobel Prize in Biomedicine in 2013, this particle with a diameter of only tens of nanometers has received unprecedented attention. attention. [0003] Almost all cells secrete exosomes, and the exosomes secreted by cells carry a lot of biological information of the source cells. Therefore, exosomes are also considered as important potential biomarkers, which can reflect the physiological and biolog...

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Application Information

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Patent Type & AuthorityPatents(China)
IPC IPC(8): C12N15/10
CPCC12N15/1013C12Q2521/537C12Q2523/308C12Q2527/125
Inventor臧远胜马跃张亚楠
OwnerSHANGHAI CHANGZHENG HOSPITAL