HIGH-DENSITY HYBRIDOMA PERFUSION CULTURE - LIMITATION VS INHIBITION

Because our earlier work indicated a strong correlation between specif ic antibody productivity and cell density in perfusion culture, we con ducted experiments to determine the optimum means of increasing cell d ensity while mainitaining high antibody productivity. The rates of med ium supply and waste removal were varied to determine whether cell den sity was limited or inhibited, and whether a diffusable substance coul d be responsible for the correlation between antibody productivity and cell density. Nutrient supply was found to be a stronger determinant of cell density than waste removal; however, the rate of waste removal had a greater effect on cell growth at lower cell densities. Even at noninhibitory levels of ammonia and lactate, cellular metabolism was r egulated to minimize their concentrations at lowered rates of waste re moval. Separate step changes in glucose and glutamine resulted in incr eased cell density and antibody concentration. Specific antibody produ ctivity increased following the step in glutamine, but not glucose. Bo th steps caused changes in cellular metabolism that prevented the leve ls of lactate and ammonia from reaching toxic levels.