Abstract
Perfusion of cultures of suspended recombinant Chinese hamster ovary (CHO) cells was accomplished through the use of an external, 3-port inclined settler which was shown to preferentially retain viable cells in the bioreactor. By vibrating the settler and cooling it to 4°C, an average of 89% of viable cells were returned to the bioreactor while the return of nonviable cells was only 72%, with the rejected cell population being only 36% viable when settler feed contained 78% viable cells. The residence time of CHO cells in the cooled, vibrated settler was determined by flow cytometric discrimination of tracer recombinant CHO cells. Cells returning to the bioreactor through the underflow had an average residence time of 1.46 hours in the settler. During perfusion cultures with cell densities above 106 cells/mL, cells seen to be stalled within the settler were easily dislodged by periodic air bubbling using a simple backflushing procedure in which headspace gas was brought through the settler underflow port. 87% of the nonretuming cells, which represented about 25% of the cells which settled, were returned to the bioreactor after bubbling within an average of 32 minutes after bubbling.
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