Single cell analysis applied to antibody fragment production with Bacillus megaterium: development of advanced physiology and bioprocess state estimation tools

2.50
Hdl Handle:
http://hdl.handle.net/10033/620759
Title:
Single cell analysis applied to antibody fragment production with Bacillus megaterium: development of advanced physiology and bioprocess state estimation tools
Authors:
David, Florian; Berger, Antje; Hänsch, Robert; Rohde, Manfred ( 0000-0003-0522-3580 ) ; Franco-Lara, Ezequiel
Abstract:
Abstract Background Single cell analysis for bioprocess monitoring is an important tool to gain deeper insights into particular cell behavior and population dynamics of production processes and can be very useful for discrimination of the real bottleneck between product biosynthesis and secretion, respectively. Results Here different dyes for viability estimation considering membrane potential (DiOC2(3), DiBAC4(3), DiOC6(3)) and cell integrity (DiBAC4(3)/PI, Syto9/PI) were successfully evaluated for Bacillus megaterium cell characterization. It was possible to establish an appropriate assay to measure the production intensities of single cells revealing certain product secretion dynamics. Methods were tested regarding their sensitivity by evaluating fluorescence surface density and fluorescent specific concentration in relation to the electronic cell volume. The assays established were applied at different stages of a bioprocess where the antibody fragment D1.3 scFv production and secretion by B. megaterium was studied. Conclusions It was possible to distinguish between live, metabolic active, depolarized, dormant, and dead cells and to discriminate between high and low productive cells. The methods were shown to be suitable tools for process monitoring at single cell level allowing a better process understanding, increasing robustness and forming a firm basis for physiology-based analysis and optimization with the general application for bioprocess development.
Citation:
Microbial Cell Factories. 2011 Apr 15;10(1):23
Issue Date:
15-Apr-2011
URI:
http://dx.doi.org/10.1186/1475-2859-10-23; http://hdl.handle.net/10033/620759
Type:
Journal Article
Appears in Collections:
publications of the central unit for microscopy (ZEIM)

Full metadata record

DC FieldValue Language
dc.contributor.authorDavid, Florianen
dc.contributor.authorBerger, Antjeen
dc.contributor.authorHänsch, Roberten
dc.contributor.authorRohde, Manfreden
dc.contributor.authorFranco-Lara, Ezequielen
dc.date.accessioned2017-01-27T08:52:31Z-
dc.date.available2017-01-27T08:52:31Z-
dc.date.issued2011-04-15en
dc.identifier.citationMicrobial Cell Factories. 2011 Apr 15;10(1):23en
dc.identifier.urihttp://dx.doi.org/10.1186/1475-2859-10-23en
dc.identifier.urihttp://hdl.handle.net/10033/620759-
dc.description.abstractAbstract Background Single cell analysis for bioprocess monitoring is an important tool to gain deeper insights into particular cell behavior and population dynamics of production processes and can be very useful for discrimination of the real bottleneck between product biosynthesis and secretion, respectively. Results Here different dyes for viability estimation considering membrane potential (DiOC2(3), DiBAC4(3), DiOC6(3)) and cell integrity (DiBAC4(3)/PI, Syto9/PI) were successfully evaluated for Bacillus megaterium cell characterization. It was possible to establish an appropriate assay to measure the production intensities of single cells revealing certain product secretion dynamics. Methods were tested regarding their sensitivity by evaluating fluorescence surface density and fluorescent specific concentration in relation to the electronic cell volume. The assays established were applied at different stages of a bioprocess where the antibody fragment D1.3 scFv production and secretion by B. megaterium was studied. Conclusions It was possible to distinguish between live, metabolic active, depolarized, dormant, and dead cells and to discriminate between high and low productive cells. The methods were shown to be suitable tools for process monitoring at single cell level allowing a better process understanding, increasing robustness and forming a firm basis for physiology-based analysis and optimization with the general application for bioprocess development.en
dc.titleSingle cell analysis applied to antibody fragment production with Bacillus megaterium: development of advanced physiology and bioprocess state estimation toolsen
dc.typeJournal Articleen
dc.language.rfc3066enen
dc.rights.holderDavid et al; licensee BioMed Central Ltd.en
dc.date.updated2015-09-04T08:28:55Zen
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