Department of Cell Engineering, Institute of Biotechnology, 20 Dongdajie, Fengtai, Beijing, 100071, P.R. China,
Cytotechnology. 2000 Jul;33(1-3):13-9. doi: 10.1023/A:1008127310890.
A recombinant DNA CHO cell line secretingurokinase-type plasminogen activator (u-PA) wascultivated with Cytopore cellulose porousmicrocarriers in a 30l Biostat UC stirred tankreactor. After 26 days of culture, using a spinfilter toretain cells in bioreactor, the cell density couldreach 1.33 x 10(7) ml(-1). The maximal u-PAactivity in supernatant was 7335 IU.ml(-1), and204l supernatant containing 7.1 g u-PA was harvested.After 100 days of culture with 0.1% fetal bovineserum medium, a modified cell retention system whichcan be washed-out backward, substituted thespinfilter to prevent filter clogging. The maximalcell density was over 10(7) ml(-1), the maximalu-PA activity in supernatant reached 6250IU.ml(-1), and 1604l supernatant containing about51 g u-PA was harvested. Compared to perfusionculture, batch medium-replaced culture could raiseutilizing efficiency of the medium, increase cell specificproductivity and improve the quality of the product which wasnot steady in a 37 degrees C environment. Cells can movefrom seed porous microcarriers occupied by cells tovacant microcarriers spontaneously, withouttrypsinization, and continue to grow until all microcarriers contained cells. It shows that Cytoporeporous microcarriers are very useful and convenient toscale up cultivation step by step.
用 Cytopore 纤维素多孔微载体在 30L Biostat UC 搅拌罐式生物反应器中培养分泌尿激酶型纤溶酶原激活剂(u-PA)的重组 DNA CHO 细胞系。培养 26 天后，使用旋转过滤器在生物反应器中保留细胞，细胞密度可达到 1.33×10(7)ml(-1)。上清液中 u-PA 的最大活性为 7335IU.ml(-1)，收获了含有 7.1g u-PA 的 204l 上清液。经过 100 天用含 0.1%胎牛血清的培养基培养后，用可反向冲洗的改良细胞保留系统代替旋转过滤器，以防止过滤器堵塞。细胞密度超过 10(7)ml(-1)，上清液中 u-PA 的最大活性达到 6250IU.ml(-1)，收获了含有约 51g u-PA 的 1604l 上清液。与灌注培养相比，分批更换培养基的培养可以提高培养基的利用率，增加细胞比生产率，并改善产品质量，因为产品在 37°C 环境中不稳定。细胞可以从被细胞占据的种子多孔微载体自发地转移到空的微载体上，而无需使用胰蛋白酶消化，并继续生长，直到所有微载体都充满细胞。这表明 Cytopore 多孔微载体非常有用且方便，可以逐步放大培养步骤。
