Cell large-scale culture method based on porous nanoscale temperature-sensitive soft colloid

Abstract

The invention belongs to the technical field of biomedical materials, and particularly relates to a cell large-scale culture method based on porous nanoscale temperature-sensitive soft colloid. The method comprises the following steps: mixing the porous nanoscale temperature-sensitive soft colloid with a cell or tissue culture to obtain a mixture, conveying the mixture in a liquid state into a hollow column reactor, and performing curing at 32-37 DEG C for culture. According to the culture method, pancreatin digestion is not needed in the cell elution process, cells are not affected at all, and the cells can be conveniently amplified into reactors of more levels. Generally, the density of the cells after first-stage amplification can reach 108 per milliliter, the cells carried by the colloid are digested together through temperature changes, a previous inoculation method is repeated to input the cells into other reactors to realize amplification, or the diameter of a hollow column cavity is increased to realize volume expansion, and process amplification and culture volume amplification are realized by increasing the number of hollow fiber columns.

Description

technical field [0001] The invention belongs to the technical field of biomedical materials, and specifically relates to a method for large-scale cell culture based on porous nanoscale temperature-sensitive soft colloids and hollow columns, which can cultivate animal stem cells or other tissue cells in a relatively large area in 3D. Background technique [0002] With the development of modern medicine, there is an increasing demand for large-scale expansion of animal stem cells or other tissue cells. The biopharmaceutical industry using CHO cells to express monoclonal antibodies has made the process of suspension culture cells very mature and achieved commercial production of tens of thousands of liters. With the advancement of technology, the equipment for cultivating CHO cells has been continuously expanded to the level of 10,000 liters. But these systems have their limitations. For example, most adherent cells cannot be directly suspended in reactor volumes above 10 L. ...

AI Technical Summary

Problems solved by technology

Adherent cell culture is generally carried out in square flask culture in a conventional laboratory or by using a commercial multi-layer cell factory. However, on the one hand, since the cells only grow on one plane, the usable area for culture is very limited, often within the range of one square centimeter. only 10 4 Level cells, if you want to reach 10 9 -10 10 The number of cells at the level requires a large number of cell factories or cell plate stacks, requiring a lot of manual and testing work, and once infected with other viruses or microorganisms, it will be fatal to the patient

In addition, plate culture belongs to 2D culture, and there is no 3D tissue structure similar to that in animals. Therefore, many studies have shown that under 2D culture conditions, many stem cells do not have the biological characteristic markers cultured in animals, and there are also some gene expressions. A large difference will greatly reduce the effect of cell therapy

The second traditional culture method is suspension culture, such as roller bottles, shake flasks, or small shear-controlled bioreactors like Wave reactors. The advantage of this method is that it greatly expands the three-dimensional culture. space, so its cell density can be greatly increased, but because suspension culture generally brings fluid shear and air bubbles to the shear damage of cell generation, the stem cells or tissues used for treatment are often primary cells. The shearing induced by air bubbles is extremely sensitive, so some scholars have pointed out that cells grown in this environment will also lose some biomarkers or show differences in gene expression

The biggest problem is that it still needs to use trypsin or other analogues for elution, which greatly affects the activity of cells and the possibility of process scale-up

Secondly, the surface of the hollow fiber is still a 2D environment. Although many newer designs have optimized the surface of the hollow fiber column to become porous and other structures, it is still difficult to simulate the 3D tissue environment, which will affect the expression of characteristic proteins on the cell surface. and gene-level expression in whole cells

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