title: Cell death in bioreactors: a role for apoptosis.
abstract: The incidence of apoptotic and necrotic cell death was compared in CHO, SF9 insect cells and murine plasmacytoma (J558L) and hybridoma (TB/C3) cells during in vitro cultivation in batch cultures. Acridine orange staining and fluorescence microscopy enabled the visualization of a classic morphological feature of apoptotic cell, the presence of condensed and/or fragmented chromatin. DNA gel electrophoresis was employed to show an additional characteristic of the process, the endonuclease-mediated fragmentation of DNA into multiples of 180 base pairs. The levels of apoptosis at the end of batch cultures of plasmacytoma and hybridoma cell lines were found to be 60% and 90% of total dead cells, respectively. However, employing the above-mentioned techniques, the biochemical and morphological features of apoptosis were not found in CHO and SF9 insect cells. Some factors affecting the induction of apoptosis during the batch culture of the hybridoma and plasmacytoma cell lines were identified. The most effective inducer was found to be glutamine limitation, followed by (in order of importance) serum limitation, glucose limitation, and ammonia toxicity. Blockage of the cell cycle of the plasmacytoma and hybridoma cells using thymidine resulted in the induction of apoptosis. This has important implications for the development of cell culture processes that minimize cell division and thereby increase specific productivity.
authors.affiliation: BBSRC Centre for Biochemical Engineering, School of Chemical Engineering, The University of Birmingham, Birmingham B15 2TT, United Kingdom.
authors.forename: R P
authors.initials: RP
authors.lastname: Singh
authors.forename: M
authors.initials: M
authors.lastname: Al-Rubeai
authors.forename: C D
authors.initials: CD
authors.lastname: Gregory
authors.forename: A N
authors.initials: AN
authors.lastname: Emery