Continuous cell propagation using low-charge microcarriers

Microcarrier cell culture technology has been extended by the finding that two mammalian epithelial cell lines can be continuously subcultured by simple bead-to-bead transfer in normal medium in which calcium concentrations have been reduced. Data are reported which show that the hamster ovary line CHO-Kl and the monkey kidney line LLC-MK₂ can be subcultured simply by adding fresh microcarriers to the stirred suspension culture. Thirteen generations of continuous exponential growth are demonstrated with two such subcultures for the CHO-Kl cells and with four such subcultures for the LLC-MK₂ cells. Cell generation times were unchanged by this subculturing approach compared to standard subculturing procedure using trypsin to remove cells from surfaces. We have applied this technique to the production of vesicular stomatitis virus (VSV) from CHO-Kl cells. Viral yields were comparable (less than twofold difference) in microcarrier cultures which were subcultured via bead-to-bead transfer or by the standard means of removing cells from microcarriers with trypsin.