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Cryopreservation solution of tissue engineering products and application method thereof

A cryopreservation and tissue engineering technology, applied in the field of tissue engineering of biomedical materials, can solve the problems of low cryopreservation efficiency, decline in the effect of tissue engineering products, and unstable structure, and achieve long storage time, low toxicity, and wide application range Effect

Inactive Publication Date: 2010-06-09
FOURTH MILITARY MEDICAL UNIVERSITY +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the efficiency of such cryopreservation is still low, and its effect will be further reduced when applied to tissue engineering products with relatively unstable structures.

Method used

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  • Cryopreservation solution of tissue engineering products and application method thereof
  • Cryopreservation solution of tissue engineering products and application method thereof
  • Cryopreservation solution of tissue engineering products and application method thereof

Examples

Experimental program
Comparison scheme
Effect test

example 1

[0017] Example 1. Cryopreservation of tissue engineered skin

[0018] 1. Preparation and pretreatment of cryopreservation solution: The cryopreservation solution uses DMEM medium as the solvent, and is added with: vitamin B 6μg / ml, vitamin C 6μg / ml, chondroitin sulfate 3% w / v, β-integration 0.4% w / v, cromolyn sodium 4μg / ml, cytochalasin B 3μg / ml, L-glutamine 4mM, bovine serum albumin 2% w / v, fetal bovine serum 10% w / v, Na 2 CO 3 0.3% w / v, trehalose 3% w / v, Ficoll-70 2% w / v;

[0019] Put the tissue engineered skin into a freeze-resistant container (such as a polyethylene or polypropylene plastic container), add the preservation solution to immerse the skin, and shake at 100 revolutions / min for 50 minutes at room temperature; add dimethyl sulfoxide to the preservation solution to make When the concentration reaches 5%, let it stand for 10 minutes after mixing; cool the preservation solution containing the skin to 4°C, add dimethyl sulfoxide to the preservation solution again to make ...

example 2

[0021] Example 2. Cryopreservation of tissue engineered blood vessels

[0022] 1. Preparation and pretreatment of cryopreservation solution: The cryopreservation solution uses DMEM medium as the solvent, and it is added with: vitamin B 15μg / ml, vitamin C 15μg / ml, chondroitin sulfate 5% w / v, β-integration 0.4% w / v, cromolyn sodium 2μg / ml, cytochalasin B 1.5μg / ml, L-glutamine 4mM, bovine serum albumin 3% w / v, fetal bovine serum 10% w / v, Na 2 CO 3 0.2% w / v, trehalose 3% w / v, Ficoll-70 2% w / v.

[0023] Place the blood vessel in a freeze-resistant container, add the protection solution to immerse the blood vessel, and shake at 80 revolutions / min for 30 minutes at room temperature; add dimethyl sulfoxide to the protection solution to make the concentration reach 3%, mix well, and stand still 10 minutes; cool down to 4℃, then add dimethyl sulfoxide to the protection solution to make the concentration reach 6%, mix well and let stand for 10 minutes; after cooling the protection solution t...

example 3

[0025] Example 3. Cryopreservation of tissue engineered cornea

[0026] 1. Preparation and pretreatment of cryopreservation solution: The cryopreservation solution uses DMEM medium as the solvent, which is added with: vitamin B 10μg / ml, vitamin C 10μg / ml, chondroitin sulfate 5% w / v, β-integration 0.4% w / v, cromolyn sodium 3μg / ml, cytochalasin B 5μg / ml, L-glutamine 4mM, bovine serum albumin 3% w / v, fetal bovine serum 10% w / v, Na 2 CO 3 0.2% w / v, trehalose 6% w / v, Ficoll-70 3% w / v;

[0027] Put the tissue engineered cornea into a freeze-resistant container, add the protective solution to immerse the cornea, and shake at 60 rpm for 40 minutes at room temperature; then add dimethyl sulfoxide to the protective solution to make the concentration reach 2%, and mix well Then let it stand for 20 minutes; lower the temperature to 4°C, add dimethyl sulfoxide again to make the concentration reach 4%, mix well and let stand for 10 minutes; after cooling down to 0°C, add dimethyl sulfoxide to m...

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Abstract

A cryopreservation solution of tissue engineering products uses DMEM culture solution as a basic solvent which is added with vitamin B, vitamin C, chondroitin sulfate, beta-integrin, cromolyn sodium, cytochalasin B, L-glutamine, bovine serum albumin, fetal bovine serum, trehalose, sodium carbonate, polysucrose-70, and dimethyl sulfoxide added during freezing storage. The cryopreservation solution provided by the invention has little toxicity to cells and long storage time, and can be stored for six months under 80 DEG C below zero, and 12 to 18 months in liquid nitrogen; and the cell viability of the resuscitated cells can be over 60%. The stored tissues can be simply and conveniently used after being resuscitated for only three to five min in water bath under the temperature of 37 DEG C and being washed by sterilized saline water. The invention can be widely used, and is suitable for tissue engineering skin, tissue engineering cornea, tissue engineering blood vessel, tissue engineering nerve and the like, and also is applicable to the cryopreservation of normal tissues.

Description

Technical field [0001] The invention belongs to the technical field of tissue engineering of biomedical materials, and specifically relates to a cryopreservation method of tissue engineering products. Background technique [0002] Tissue engineering products are artificial tissues or organs constructed from cells and three-dimensional scaffolds. For example, tissue-engineered skin constructed in vitro is a functional skin substitute constructed by combining seed cells with scaffold materials, and then planting them on In wounds, in the process of gradual degradation and absorption of scaffold materials, cells can grow, differentiate and form tissues with certain functions, so as to replace, repair, and regenerate damaged tissues or organs. The ultimate goal of tissue engineering is to achieve large-scale industrialization, so the long-term preservation of tissue engineering products is an indispensable and important link. [0003] Since tissue engineering products contain living c...

Claims

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Application Information

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IPC IPC(8): A01N1/02
Inventor 金岩罗海浪张勇杰
Owner FOURTH MILITARY MEDICAL UNIVERSITY
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