Abstract
The death of the hybridoma VO 208 in a continuous culture at pH 7 and 6.8 was investigated by measuring both the appearance of visible dead cells which do not exclude the trypan blue dye and the release of lactate dehydrogenase (LDH) in the culture medium. The intracellular LDH was found to be completely released either when live cells lysed or when they were transformed into visible dead cells. No significant lysis of blue dead cells could be observed at the two different pH. Using a LDH balance over the culture system, cell lysis was found negligible at pH 7, but accounted for 20% of the total cell death at pH 6.8. A methodology is proposed to evaluate the rate constants of hybridoma lysis and total death. For the investigated cell line in continuous culture, the calculated total cell death rate constant was found to increase from 0.002 h−1 to 0.01 h−1 when decreasing the pH from 7 to 6.8.
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Abbreviations
- D:
-
dilution rate (h−1)
- kb :
-
specific trypan-blue dead cells appearance rate (h−1)
- kL :
-
specific lysis rate of viable cells (h−1)
- kd :
-
specific death rate (h-1)
- LDH0 :
-
lactate dehydrogenase activity in the feed culture medium (IU.l−1)
- LDH:
-
lactate dehydrogenase activity in the outlet culture medium (IU.l−1)
- LDHi :
-
intracellular lactate dehydrogenase activity of viable cells (IU.10−9 cells)
- rLDH :
-
total rate of LDH release (IU.h−1.L−1)
- rb :
-
transformation rate of viable cells into blue dead cells (109 cells.h−1.L−1)
- xv :
-
viable cell concentration (109 cells.l−1)
- xb :
-
trypan-blue dead cell concentration (109 cells.l−1)
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Goergen, J.L., Marc, A. & Engasser, J.M. Determination of cell lysis and death kinetics in continuous hybridoma cultures from the measurement of lactate dehydrogenase release. Cytotechnology 11, 189–195 (1993). https://doi.org/10.1007/BF00749869
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DOI: https://doi.org/10.1007/BF00749869