An Advanced Monitoring Platform for Rational Design of Recombinant Processes - Measurement, Monitoring, Modelling, and Control of Bioprocesses - page 70

cannot be fully exploited, resulting in too low yield and noncompliant product
quality.
During evolution, cellular systems have developed a multitude of mechanisms
to cope with a broad range of stress situations, and a complex regulatory network
guarantees optimal response reactions [ 32 ]. Stress triggered by recombinant gene
expression originates in the cell, and the recombinant system is not or only partly
controlled by the cellular regulatory network. In consequence, countermeasures of
the cellular stress response system are not adequate and often inefficient [ 30 , 31 ].
The main challenge in host cell design is that understanding of the biology of
protein expression is still fragmentary and incomplete.

2 A Process Monitoring Platform for Recombinant
Protein Production

The configuration of the established platform for monitoring of recombinant
protein production processes with E. coli as the host cell is designed according to
the demands of these processes. Beside routine monitoring of physical (e.g.,
agitator speed, flow rates, and temperature) and chemical variables (e.g., pH, pO 2 ,
and CO 2 and O 2 in the off-gas), a series of state-of-the-art bioprocess monitoring
devices are implemented in the established online monitoring platform. Online
techniques are combined with accurate offline methods to allow for monitoring and
measurement of cell growth and cell viability, quantification of recombinant
compounds, and characterization of the cellular response to recombinant gene
expression. This section describes how we developed such a platform for

recombinant protein expression in E. coli.

2.1 Monitoring of Biomass and Growth

Biomass, viability, and growth kinetics are among the most critical bioprocess
variables and therefore attractive monitoring targets for the process monitoring
platform. The classical offline methods for determination of biomass are optical
density measurements and gravimetric cell dry mass quantification. In face of the
importance of these variables, development of methods for in situ measurement
was a focus of the research [ 33 - 35 ]. Despite all these efforts and achievements,
real-time measurement remains challenging.
Optical density probes are the most commonly used in situ devices for online
biomass estimation. The TruCell TM cell growth monitor (Finesse Solutions AG,
Bonaduz, Switzerland), an optical transmission sensor that enables in situ
monitoring of fermentation processes, was selected from among the available
optical probes.