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Kit for separating exosome from cell supernatant and use method of kit

A technology for supernatant and exosomes, which is applied in the field of kits for separating exosomes, can solve the problems of inability to be widely used, and achieve the effects of easy operation, high reliability and high purity

Inactive Publication Date: 2021-08-06
WUHAN LIFE ORIGIN BIOTECH LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, considering the particularity of the required equipment, it cannot be widely used

Method used

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  • Kit for separating exosome from cell supernatant and use method of kit
  • Kit for separating exosome from cell supernatant and use method of kit
  • Kit for separating exosome from cell supernatant and use method of kit

Examples

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Embodiment 1

[0087] Example 1: The kit for isolating exosomes from cell supernatant according to the present invention and its application method

[0088] The kit of the present invention comprises reagent A, reagent B, reagent C and reagent D;

[0089] Wherein, the preparation step of the reagent A is: mixing deionized water and PEG 6000 (protein precipitation agent), so that the concentration of the reagent A is 35g / mL, and then performing sterilization treatment;

[0090] Described reagent B is Protein A / G (BeaverBeads TM , No. 22202-100) solution;

[0091] The preparation steps of the reagent C are as follows: mixing deionized water and antibody CD63 so that the concentration of the obtained antibody CD63 solution is 20 μg / mL, and then performing sterilization treatment;

[0092] The preparation step of the reagent D is: mixing PBS and 0.05% Tween 20; wherein the preparation method of the PBS is: mixing 7g NaCl, 0.1g KCl, 1g Na 2 HPO 4 and 0.2g KH 2 PO 4 Mix, add water to 0.7L;...

Embodiment 2

[0107] Example 2: The kit for isolating exosomes from cell supernatant according to the present invention and its application method

[0108] The kit of the present invention includes reagent A, reagent D and reagent G;

[0109] Wherein, the preparation step of the reagent A is: mixing deionized water and PEG 6000 (protein precipitation agent), so that the concentration of the reagent A is 35g / mL, and then performing sterilization treatment;

[0110] The preparation step of the reagent D is: mixing PBS and 0.05% Tween 20; wherein the preparation method of the PBS is: mixing 7g NaCl, 0.1g KCl, 1g Na 2 HPO 4 and 0.2g KH 2 PO 4 Mix, add water to 0.7L;

[0111] The reagent G is a solution of magnetic particles coated with CD63 at a concentration of 0.5-10%, and the preparation steps of the reagent G are as follows: take the magnetic particle Protein A / G (BeaverBeads TM , No. 22202-100) solution 10μl, separated for 2min, discarded the supernatant to obtain the first solution...

Embodiment 3

[0126] Example 3: The kit for isolating exosomes from cell supernatant according to the present invention and its application method

[0127] The kit of the present invention comprises reagent A, reagent B, reagent C and reagent D;

[0128] Wherein, the preparation step of the reagent A is: mixing deionized water and PEG 8000 (protein precipitation agent), so that the concentration of the reagent A is 45g / mL, and then performing sterilization treatment;

[0129] The reagent B is magnetic particle Protein A / G (BeaverBeads TM , No. 22202-100) solution;

[0130] The preparation steps of the reagent C are as follows: mixing deionized water and antibody CD81 so that the concentration of the obtained antibody CD81 solution is 100 μg / mL, and then performing sterilization treatment;

[0131] The preparation step of the reagent D is: mix PBS and 1% Tween 20; wherein the preparation method of the PBS is: mix 9g NaCl, 0.3g KCl, 2g Na 2 HPO 4 and 1g KH 2 PO 4 Mix, add water to 1.5...

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Abstract

The invention provides a kit for separating exosome from cell supernate. The kit comprises a reagent A, a reagent B, a reagent C and a reagent D, wherein the reagent A is a PEG 6000 and / or PEG 8000 aqueous solution with the concentration of 35 to 45 g / mL; the reagent B is a magnetic particle solution with the concentration of 1-10%; the reagent C is an antibody solution with the concentration of 20-100 [mu] g / mL, wherein an antibody is selected from one or more than two of CD63, CD81 and CD9; and the reagent D is a Tween 20 phosphate buffer solution with the concentration of 0.05 to 1%. According to the kit, the use method and the application, the high-purity exosome can be efficiently obtained, the influence of substances such as impure protein and the like is eliminated, and the kit has the characteristics of simplicity, convenience, stability, high purity, easiness in operation, high reliability in analysis of protein components and the like.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a kit for isolating exosomes from cell supernatants and a method for using the same. Background technique [0002] Exosomes are subcellular vesicle structures secreted by various cells with a diameter of 30-140 nm, which appear spherical or cup-shaped under an electron microscope. Exosomes originate from early endosomes in cells, which bud inwardly to form multivesicular endosomes (mμltivesicμlar endosomes), which fuse with the cell membrane to release small vesicles outside the cell. Exosomes are formed. The component analysis of exosomes shows that exosomes contain a large number of proteins, nucleic acids and lipids, some of which are common components common to all exosomes, and the rest are unique due to different exosome source tissues components. No matter which component it is, it plays a key role in the process of exosome-mediated intercellular material and in...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/00
CPCC12N5/00C12N2509/00G01N1/40C12N5/06G01N1/28G01N1/34
Inventor 于倩华权高沈鹤霄易汪雪蔡赢柯红郭静舒芹
Owner WUHAN LIFE ORIGIN BIOTECH LTD
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