KINETICS AND STABILITY OF GM-CSF PRODUCTION BY RECOMBINANT YEAST-CELLS IMMOBILIZED IN A FIBROUS-BED BIOREACTOR

The continuous production of murine granulocyte-macrophage colony-stim ulating factor (GM-CSF) by recombinant yeast cells immobilized in a fi brous-bed bioreactor was studied. A high cell density of similar to 68 g/L and a GM-CSF productivity of similar to 3.5 mg/ L . h were attain ed in the fibrous-bed bioreactor fed with a rich (nonselective, pH 6.7 ) medium at a dilution rate of 0.16 h(-1). The GM-CSF production was s table even though the fraction of plasmid-carrying cells in the reacto r effluent gradually dropped below 5% over a period of 2 weeks. At the end of that period, the immobilized cells in the fibrous matrix still had a high fraction, similar to 26%, of plasmid-carrying cells. Simil ar results were obtained with reactors operated at 0.05 h(-1) dilution rate and pH 4.0. Although the GM-CSF production was lower at pH 4, th e reactor was stably operated for over 4 weeks without contamination o r significant loss of productivity. The stable longterm GM-CSF product ion from the fibrous-bed bioreactor was attributed to the effect of ce ll immobilization on plasmid stability. Because GM-CSF production was growth-associated, as was found in batch fermentation with free cells, this stabilization effect cannot be attributed solely to the reduced cell growth in the immobilized cell environment. Plasmid-carrying cell s were preferentially retained in the fibrous matrix, perhaps because their abilities to adhere to the fiber surface and to form cell aggreg ates were higher than those of plasmid-free cells.