The use of multi-parameter flow cytometry to compare the physiological response of Escherichia coli W3110 to glucose limitation during batch, fed-batch and continuous culture cultivations

Hewitt, C.J.; Nebe-von Caron, G.; Nienow, A.W.; McFarlane, C.M.

Journal of Biotechnology 75(2-3): 251-264


ISSN/ISBN: 0168-1656
PMID: 10553662
DOI: 10.1016/s0168-1656(99)00168-6
Accession: 011558263

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Multi-parameter flow cytometric techniques have been developed for the 'at-line' study of bacterial cultivations. Using a mixture of specific fluorescent stains it is possible to resolve an individual cells physiological state beyond culturability, based on the presence or absence of an intact polarised cytoplasmic membrane, enabling assessment of population heterogeneity. It has been shown that during the latter stages of small-scale (5 l), well mixed fed-batch cultivations there is a considerable drop in cell viability, about 17%, as characterised by cytoplasmic membrane depolarisation and permeability. These phenomena are thought to be due to the severe and steadily increasing stress associated with glucose limitation at high cell densities, during the fed-batch process. Such effects were not found in either batch or continuous culture cultivations. The possibility of using these findings for improved process control using 'on-line' flow cytometry are discussed.