Pilot production of u-PA with porous microcarrier cell culture

A recombinant DNA CHO cell line secreting urokinase-type plasminogen activa tor (u-PA) was cultivated with Cytopore cellulose porous microcarriers in a 30l Biostat UC stirred tank reactor. After 26 days of culture, using a spi nfilter to retain cells in bioreactor, the cell density could reach 1.33 x 10(7) ml(-1). The maximal u-PA activity in supernatant was 7335 IU.ml(-1), and 204l supernatant containing 7.1 g u-PA was harvested. After 100 days of culture with 0.1% fetal bovine serum medium, a modified cell retention sys tem which can be washed-out backward, substituted the spinfilter to prevent filter clogging. The maximal cell density was over 10(7) ml(-1), the maxim al u-PA activity in supernatant reached 6250 IU.ml(-1), and 1604l supernata nt containing about 51 g u-PA was harvested. Compared to perfusion culture, batch medium-replaced culture could raise utilizing efficiency of the medi um, increase cell specific productivity and improve the quality of the prod uct which was not steady in a 37 degrees C environment. Cells can move from seed porous microcarriers occupied by cells to vacant microcarriers sponta neously, without trypsinization, and continue to grow until all microcarrie rs contained cells. It shows that Cytopore porous microcarriers are very us eful and convenient to scale up cultivation step by step.