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Method for three-dimensional culture of fibrous ring derived stem cells based on Collagel gel bracket method

A technology of three-dimensional culture and annulus fibrosus, applied in the fields of biotechnology and cell culture, can solve the problems of unsatisfactory effect of three-dimensional cell culture technology, and achieve the effect of ensuring tissue moisture, enhancing adhesion ability, and not easy to fall off

Inactive Publication Date: 2020-09-25
SHANDONG MEDICAL BIO TECH RES CENT
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] For this reason, the technical problem to be solved by the present invention is to provide a method for three-dimensional culture of annulus fibrosus stem cells based on the Collagel gel scaffold method, so as to solve the unsatisfactory effect of traditional three-dimensional cell culture technology in the prior art

Method used

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  • Method for three-dimensional culture of fibrous ring derived stem cells based on Collagel gel bracket method
  • Method for three-dimensional culture of fibrous ring derived stem cells based on Collagel gel bracket method
  • Method for three-dimensional culture of fibrous ring derived stem cells based on Collagel gel bracket method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0044] The Collagel culture medium matrix described in the present embodiment comprises following composition:

[0045] Reagent A: 5×MEM, 288 μl;

[0046] Reagent B: FBS, 144 μl;

[0047] Reagent C: Hepes (1M), 36 μl;

[0048] Reagent D: 0.1M NaOH and 5×MEM mixture, 72μl;

[0049] Collagel: 3mg / ml COL1 mixed with 0.01M HCl, 1260μl.

[0050] Take 288 μl of Reagent A (5×MEM), 144 μl of Reagent B (FBS) and 36 μl of Reagent C (Hepes, 1M) in Test Tube 1 for thorough mixing and set aside; another 1260 μl of Collagel (3mg / ml COL1 mixed with 0.01M HCl solution) and 72 μl of reagent D (mixed solution of 0.1M NaOH and 5×MEM) were placed in test tube 2 for thorough mixing, and the pH of the mixed solution was adjusted to 7.0 for later use; the mixed solutions in test tube 1 and test tube 2 were mixed to obtain the obtained Need Collagel culture substrate, spare. It should be noted that the storage and operation of all the above reagents need to be kept at 4°C or on ice.

[0051] Se...

Embodiment 2

[0056] The Collagel culture medium matrix described in the present embodiment comprises following composition:

[0057] Reagent A: 5×MEM, 288 μl;

[0058] Reagent B: FBS, 144 μl;

[0059] Reagent C: Hepes (1M), 36 μl;

[0060] Reagent D: 0.1M NaOH and 5×MEM mixture, 72μl;

[0061] Collagel: 3mg / ml COL1 mixed with 0.01M HCl, 1260μl.

[0062] Take 288 μl of Reagent A (5×MEM), 144 μl of Reagent B (FBS) and 36 μl of Reagent C (Hepes, 1M) in Test Tube 1 for thorough mixing and set aside; another 1260 μl of Collagel (3mg / ml COL1 mixed with 0.01M HCl solution) and 72 μl of reagent D (mixed solution of 0.1M NaOH and 5×MEM) were placed in test tube 2 for thorough mixing, and the pH of the mixed solution was adjusted to 7.0-7.2 for later use; mix the mixed solutions in test tube 1 and test tube 2, Obtain the required Collagel culture substrate for later use. It should be noted that the storage and operation of all the above reagents need to be kept at 4°C or on ice.

[0063] Selec...

Embodiment 3

[0067] The Collagel culture medium matrix described in the present embodiment comprises following composition:

[0068] Reagent A: 5×MEM, 288 μl;

[0069] Reagent B: FBS, 144 μl;

[0070] Reagent C: Hepes (1M), 36 μl;

[0071] Reagent D: 0.1M NaOH and 5×MEM mixture, 72μl;

[0072] Collagel: 3mg / ml COL1 mixed with 0.01M HCl, 1260μl.

[0073] Take 288 μl of Reagent A (5×MEM), 144 μl of Reagent B (FBS) and 36 μl of Reagent C (Hepes, 1M) in Test Tube 1 for thorough mixing and set aside; another 1260 μl of Collagel (3mg / ml COL1 mixed with 0.01M HCl solution) and 72 μl of reagent D (mixed solution of 0.1M NaOH and 5×MEM) were placed in test tube 2 for thorough mixing, and the pH of the mixed solution was adjusted to 7.0-7.2 for later use; mix the mixed solutions in test tube 1 and test tube 2, Obtain the required Collagel culture substrate for later use. It should be noted that the storage and operation of all the above reagents need to be kept at 4°C or on ice.

[0074] Selec...

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Abstract

The invention belongs to the technical field of a biological technology and cell culture, and particularly relates to a method for three-dimensional culture of fibrous ring derived stem cells based ona Collagel gel bracket method. According to the method disclosed by the invention, a Collagel gel bracket is used as a three-dimensional substrate, and the fibrous ring derived stem cells and Collagel gel are mixed to obtain cell suspension for culture, so that a tissue-like physical and space three-dimensional structure desired by cell growth can be better simulated, the microenvironment of cellgrowth under different biological mechanics actions of in vivo physiological environment can be better simulated, biological functions of growth, proliferation, adhesive attachment, disintegration and the like of the fibrous ring derived stem cells can be promoted, and the method is more suitable for research of the medical tissue project field and the biological mechanics field of intervertebraldisc tissue repair, fibrous ring tissue regeneration and the like.

Description

technical field [0001] The invention belongs to the technical field of biotechnology and cell culture, and in particular relates to a method for three-dimensionally culturing annulus fibrosus stem cells based on a Collagel gel scaffolding method. Background technique [0002] Intervertebral disc degeneration is a common disease, especially with the aging population and changes in people's work and lifestyle, the incidence of spinal degenerative diseases such as disc herniation and spinal stenosis continues to increase, and some patients with severe symptoms may even need to undergo surgery treat. At present, nucleus pulposus extraction and intervertebral fusion are the main surgical treatments for spinal degenerative diseases, but the defect of annulus fibrosus will inevitably be caused during the operation. [0003] Annulus fibrosus refers to the fibrous tissue composed of fibrous cartilage located at the periphery of the intervertebral disc. The fibers of the annulus fibr...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/0775
CPCC12N5/0668C12N2513/00C12N2500/60C12N2533/54C12N2533/52C12N2533/50
Inventor 王丹丹师彬李华忠侯冰王涛张庆浩
Owner SHANDONG MEDICAL BIO TECH RES CENT
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