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A kind of organoid culture substrate material and its preparation method and application

A technology for culturing substrate materials and organ culture, which is applied in the field of organoid culture substrate materials and their preparation, can solve the problems of large differences between batches, high prices, complex components, etc., and achieves good dryness, maintains dryness, and is simple in preparation process. Effect

Active Publication Date: 2021-11-26
ACCURATE INT BIOTECHNOLOGY (GUANGZHOU) CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, Matrigel is a very complex system, which is reflected in its complex composition, large batch-to-batch variation, and high price. Many scientific research institutions and scholars in the world are working on the research of its substitute products in order to develop a quality-reliable product. Organoid culture substrate with excellent performance and favorable price

Method used

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  • A kind of organoid culture substrate material and its preparation method and application
  • A kind of organoid culture substrate material and its preparation method and application
  • A kind of organoid culture substrate material and its preparation method and application

Examples

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Embodiment 1

[0033] This embodiment provides a method for preparing an organoid culture matrix material, comprising the following steps:

[0034] (1) Dissolving collagen type 1, collagen type 4, laminin, hyaluronic acid, and heparin with 1% acetic acid solution to obtain a collagen solution, and storing it on an ice box;

[0035] (2) Add sodium dihydrogen phosphate, EGF, FGF, A83-01, and Y27632 into the high-sugar DMEM / reduced serum DMEM mixed nutrient solution, mix well, add cells to be cultured, and then mix well with the collagen solution , adjust the pH to 7.5 with 0.1M NaOH, wherein the volume ratio of high-sugar DMEM and reduced-serum DMEM in the high-sugar DMEM / reduced serum DMEM mixed nutrient solution is 5:1, and the final concentration of each component in the cell culture medium is: Type 1 collagen 12mg / ml, laminin 3.2mg / ml, type 4 collagen 0.1mg / ml, sodium dihydrogen phosphate 1%, hyaluronic acid 1mg / ml, heparin 0.9mg / ml, A83-01 0.9ng / ml, Y27632 15uM / ml, EGF 10ng / ml, FGF 100n...

Embodiment 2

[0037] This embodiment provides a method for preparing an organoid culture matrix material, comprising the following steps:

[0038] (1) Dissolving collagen type 1, collagen type 4, laminin, hyaluronic acid, and heparin with 1% acetic acid solution to obtain a collagen solution, and storing it on an ice box;

[0039] (2) Add sodium dihydrogen phosphate, EGF, FGF, A83-01, and Y27632 into the high-sugar DMEM / serum-reduced DMEM mixed nutrient solution, mix well and then mix well with the collagen solution, and adjust the pH to 7.5 Obtained, wherein the volume ratio of high-sugar DMEM and reduced-serum DMEM in the mixed nutrient solution of high-sugar DMEM / reduced serum DMEM is 5:1, and the final concentration of each component in the cell culture medium is: collagen type 18 mg / ml, Laminin 1.5mg / ml, type IV collagen 1mg / ml, sodium dihydrogen phosphate 0.1%, hyaluronic acid 5mg / ml, heparin 0.5mg / ml, A83-01 1.9ng / ml, Y27632 8uM / ml, EGF 100ng / ml, FGF 10ng / ml.

Embodiment 3

[0041] This embodiment provides a method for preparing an organoid culture matrix material, comprising the following steps:

[0042] (1) Dissolving collagen type 1, collagen type 4, laminin, hyaluronic acid, and heparin with 1% acetic acid solution to obtain a collagen solution, and storing it on an ice box;

[0043](2) Add sodium dihydrogen phosphate, EGF, FGF, A83-01, and Y27632 into the high-sugar DMEM / serum-reduced DMEM mixed nutrient solution, mix well and then mix well with the collagen solution, and adjust the pH to 7.5 Obtained, wherein the volume ratio of high-sugar DMEM and reduced-serum DMEM in the mixed nutrient solution of high-sugar DMEM / reduced serum DMEM is 5:1, and the final concentration of each component in the cell culture medium is: collagen type 15 mg / ml, Laminin 2mg / ml, type IV collagen 0.5mg / ml, sodium dihydrogen phosphate 0.5%, hyaluronic acid 3mg / ml, heparin 0.6mg / ml, A83-01 1.2ng / ml, Y27632 10uM / ml, EGF 50ng / ml, FGF 50ng / ml.

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Abstract

The invention provides an organoid culture matrix material and its preparation method and application. The preparation method of the organoid culture matrix material comprises the following steps: mixing type I collagen, type IV collagen, laminin, hyaluronic acid, and heparin with acetic acid Dissolve the solution to obtain a collagen solution; add sodium dihydrogen phosphate, growth factors, A83-01, and Y27632 into the high-sugar DMEM / serum-reduced DMEM mixed nutrient solution, mix well and then mix with the collagen solution evenly, adjust the pH with NaOH To 7.2~8.3, vested. The organoid culture matrix material of the present invention has a cell proliferation rate higher than that of Matrigel when cultured as a matrix material in the culture of some kinds of organoids, and can better maintain the The dryness of the organ, the price is far lower than the price of Matrigel, a general-purpose organoid culture material.

Description

technical field [0001] The invention belongs to the technical field of biomedicine, and in particular relates to an organoid culture substrate material and a preparation method and application thereof. Background technique [0002] One of the key advances in stem cell research over the past decade has been the development of organoid systems. Organoids are three-dimensional (3D) cell cultures that contain some of the key properties of their representative organs. These in vitro culture systems include a population of self-renewing stem cells that can differentiate into multiple organ-specific cell types, possess similar spatial organization to, and reproduce some of the functions of, corresponding organs, thereby providing a highly physiologically relevant system . Organoids can be generated from tissue samples containing adult stem cells, from single adult stem cells, or through directed induced differentiation of pluripotent stem cells. Because some organoid model syste...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/071C12N5/09
CPCC12N5/0671C12N5/0677C12N5/0679C12N5/0688C12N5/0693C12N2500/12C12N2501/11C12N2501/115C12N2501/15C12N2501/405C12N2513/00C12N2533/52C12N2533/54C12N2533/70C12N2533/80
Inventor 黄敏尹天武徐丛廖传荣
Owner ACCURATE INT BIOTECHNOLOGY (GUANGZHOU) CO LTD
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