Microcarrier for large-scale culture cell

A large-scale culture, microcarrier technology, applied in tissue culture, tissue cell/virus culture devices, fixed on/in organic carriers, etc., can solve the problem of inability to take into account cell affinity and long-term maintenance of cell function, etc. It is suitable for large-scale culture, maintains metabolic activity, and has low cost.

Inactive Publication Date: 2007-06-13
INST OF ZOOLOGY CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The purpose of the present invention is to overcome the defect that the microcarriers of the prior art cannot take into account the affinity of cells and the long-term maintenance of cell functions, thereby providing a microcarrier that not only has the affinity of cells, but also contributes to the adhesion of cells and supports a large number of cells. Growth; and can maintain cell function for a long time, maintain cell metabolic activity and low cost microcarriers for large-scale cell culture

Method used

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  • Microcarrier for large-scale culture cell

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Experimental program
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Effect test

Embodiment 1

[0041] The preparation of embodiment 1, solid microcarrier I

[0042] Prepare 1w / v% chitosan acetic acid aqueous solution and 1w / v% gelatin aqueous solution, the two are mixed uniformly at 50 ℃ by 1: 3 volume ratio, be prepared into composite chitosan-gelatin aqueous solution; Under 300rpm mechanical stirring, Add 3 volumes of mineral oil containing 0.5w / v% Span 40 to this solution, and stir for 5 minutes to form a stable chitosan-gelatin water-in-oil agent; add 0.1w / v% pentadiene while stirring Aldehyde, continue to stir for 0.5 hours, and then stand still for 0.5 hours to obtain the aqueous phase solidified with glutaraldehyde as a crosslinking agent; filter with a 100 / 280 μm pore size sieve to collect microcarriers with a diameter of 100 to 280 μm to obtain solid microcarriers ; Wash with surfactant Tween to remove the oil phase on the surface of the carrier; then treat with 0.01M glycine for 30 hours to block unreacted aldehyde groups; then treat with 0.1w / v% NaOH for 10 h...

Embodiment 2

[0043] Embodiment 2, the preparation of solid microcarrier II

[0044] Prepare 5w / v% chitosan acetic acid aqueous solution and 5w / v% gelatin aqueous solution, the two are mixed uniformly at 70 ℃ by 3: 1 volume ratio, be prepared into composite chitosan-gelatin aqueous solution; Under 500rpm mechanical stirring, Add 10 times the volume of this solution to mineral oil containing 5w / v% Span 60, stir for 30 minutes to form a stable chitosan-gelatin water-in-oil agent; add 10w / v% glutaraldehyde while stirring, Continue to stir for 5 hours, and then stand still for 5 hours to obtain an aqueous phase solidified with glutaraldehyde as a cross-linking agent; filter through a sieve with a pore size of 100 / 280 μm, and collect microcarriers with a diameter of 100 to 280 μm to obtain solid microcarriers; Wash with petroleum ether to remove the oil phase on the surface of the carrier; then treat with 0.5M glycine for 2 hours to block unreacted aldehyde groups; then treat with 10w / v% NaOH fo...

Embodiment 3

[0045] Embodiment 3, the preparation of solid microcarrier III

[0046] Prepare 3w / v% chitosan acetic acid aqueous solution and 3w / v% gelatin aqueous solution, the two are mixed uniformly at 60 ℃ by 1: 1 volume ratio, be prepared into composite chitosan-gelatin aqueous solution; Under 400rpm mechanical stirring, Add 8 times the volume of mineral oil containing 2.5w / v% Span 85 to this solution, and stir for 20 minutes to form a stable chitosan-gelatin water-in-oil agent; add 5w / v% glutaraldehyde while stirring , continue to stir for 2 hours, and then stand still for 2 hours to obtain an aqueous phase solidified with glutaraldehyde as a crosslinking agent; filter with a sieve with a pore size of 100 / 280 μm, and collect microcarriers with a diameter of 100 to 280 μm to obtain solid microcarriers; Wash with acetone to remove the oil phase on the surface of the carrier; then treat with 0.2M glycine for 15 hours to block unreacted aldehyde groups; then treat with 4w / v% NaOH for 5 ho...

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Abstract

The present invention relates to one kind of micro carrier for large scale cell culture. The micro carrier is prepared with chitosan and gelatin and through the following steps: mixing chitosan and acetic acid aqua and gelatin aqua in certain ratio to form composite chitosan-gelatin aqua; adding mineral oil with Span to form stable chitosan-gelatin water-in-oil preparation; adding glutaraldehyde to cure; filtering with 100 / 280 micron mesh and collecting solid micro carrier, or filtering with 450 / 600 micron mesh, freeze drying and collecting porous micro carrier; processing with glycine to close un-reacted aldehyde group; and finally processing with NaOH to eliminate residual acetic acid and to obtain the micro carrier. The micro carrier is favorable to cell adhesion and can maintain cell function for long time. In addition, the method of the present invention can control various parameters of the micro carrier in relatively wide range, has low cost and is suitable for industrial production.

Description

technical field [0001] The invention relates to a microcarrier for culturing cells, in particular to a microcarrier for large-scale culturing of cells. technical background [0002] In the treatment of patients with acute and chronic liver failure caused by various reasons, the bioartificial liver support system centered on cultured hepatocytes is receiving increasing attention from the medical community. The curative effect of the bioartificial liver depends on the quantity and quality of the hepatocytes, that is, only when the number of hepatocytes cultured in vitro reaches 20-30% of the liver weight of the patient to be treated, and the cells maintain high metabolic activity in vitro, the artificial liver can be rescued. patient's life. Microcarriers are one of the supporting conditions for large-scale cell culture, and the activity and function of cells are closely related to the surface properties of microcarriers. [0003] Microcarriers made of chitosan have been use...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/02C12N11/02C12M3/00
Inventor 李克国丰美福汪蕴苗振川徐大勇
Owner INST OF ZOOLOGY CHINESE ACAD OF SCI
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