The application of multi-parameter flow cytometry to the study of recombinant Escherichia coli batch fermentation processes

Journal of Industrial Microbiology & Biotechnology
Gareth LewisC J Hewitt

Abstract

Multi-parameter flow cytometric techniques coupled with dual colour fluorescent staining were used to study the physical and metabolic consequences of inclusion body formation in batch cultures of the recombinant Escherichia coli strain MSD3735. This strain contains a plasmid coding for the isopropylthiogalactopyranoside-inducible model eukaryotic protein AP50. It is known that the synthesis of foreign proteins at high concentrations can exert a severe metabolic stress on the host cell and that morphological changes can occur. In this work, using various points of induction, it was shown that inclusion body formation is followed immediately by measurable changes in the characteristic intrinsic light scatter patterns for the individual cell (forward scatter, 90 degrees side scatter) and a concomitant progressive change in the individual cell physiological state with respect to both cytoplasmic membrane polarisation and permeability. This work establishes flow cytometry as a potentially valuable tool for monitoring recombinant fermentation processes, providing important information for scale-up. Further, we discuss the possibility of optimising inclusion body formation by manipulating the fermentation conditions based on these ra...Continue Reading

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Citations

Jun 16, 2007·Applied Microbiology and Biotechnology·C Perry Chou
Jan 1, 2008·Biotechnology Letters·Harriet LuggLynne E Macaskie
Jan 5, 2005·Microbial Cell Factories·Hans Peter Sørensen, Kim Kusk Mortensen
Sep 30, 2008·Pakistan Journal of Biological Sciences : PJBS·T HaryantiA R Raha
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Oct 1, 2008·Protein Expression and Purification·Natalia Kovalskaya, Rosemarie W Hammond
Dec 4, 2008·Cytometry. Part a : the Journal of the International Society for Analytical Cytology·Andrew WantChristopher J Hewitt
Oct 7, 2006·Biotechnology Progress·K ListnerM Chartrain
May 25, 2011·Biotechnology Progress·M A Perez-PardoD G Bracewell
Apr 26, 2016·Biotechnology Progress·Joseph M NewtonYuhong Zhou
Sep 18, 2007·Advances in Applied Microbiology·Christopher J Hewitt, Alvin W Nienow
Apr 14, 2016·Biotechnology Progress·Desmond M SchofieldDarren N Nesbeth
Mar 16, 2018·Bioprocess and Biosystems Engineering·Anna-Lena Heins, Dirk Weuster-Botz

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