in-line electrochemistry probes applied for the monitoring of the cell culture en-

vironment with the pH, pO2, and pCO2 probes are used. This allows for close

monitoring of the cellular respiration and metabolic activity (lactic acid secretion)

within the cultivation broth. Eventually some at-line tools allowing for cell counting

like automated cell counters might be used. Cell metabolites secreted or consumed

by cultivated cells could also be monitored using several technologies. To replace

the old off-line biochemical assays, novel automated metabolites analyzers were

developed and implemented at-line thanks to automated sterile sampling devices in

the last 20 years. This now allows discrete sampling of the production process to

quantify all the main cellular metabolites as well as complementary information like

trace elements or osmolarity.

Additionally, several developments were dedicated in the last 20 years to

achieving inline monitoring of viral production processes key parameters. The

technologies evaluated were mostly spectroscopic (infrared, near infrared, fluor-

escence, raman, dielectric spectroscopies) [34–37].

More specifically, if we focus on the viral production processes, we will dis-

tinguish two types of analytical tools. First, the technologies aiming at quantifying

the final product and which could be implemented at-line. Here we find again the

viral particle counter already described in section 8.3.2.2. Indeed, most viral

counters allow for viral particles count within a wide range of sample matrix from

culture supernatant up to highly purified solutions. Such assays allow for better

process understanding and monitoring of the biological events occurring in the

production phase (viral release, viral cycle length, etc.). The only information that is

not yet reached with such a technology is the activity and purity of the viral

particles. Also, information is available only at sampling times.

The second kind of technology is the one targeting the production cells. Indeed,

cells are here the production units modifications of the total cell biomass available

for production in terms of quantity or quality can strongly affect the production

efficiency. Therefore, from the late 1990s, analytical tools were developed to allow

for live biomass monitoring in cell-based production. Two examples of technolo-

gies should be emphasized here, the Raman and the capacitance spectroscopy,

besides other technologies that have also been explored.

Raman spectroscopy was originally assessed to monitor the composition of the

cultivation media over the cultivation process in cell-based processes. Indeed, this

vibrational spectroscopy tool allows for the detection of all the organic molecules. It

will record spectral fingerprints of the molecules mixture thus necessitating an

additional chemometric spectral post-treatment to extract information on the tar-

geted analytes. It was successfully applied for the monitoring of several cell

metabolites, either essential nutrients (glucose, amino acids) or metabolic cell by-

products (lactate, other acids), in cell-based production of recombinant protein

production [37–40]. Also, as cells are composed of several specific organic mole-

cules, Raman spectroscopy was concomitantly evaluated for its capacity to monitor

cell productive biomass. It was successfully applied for CHO cell culture which is

the gold standard technology for recombinant protein production [39], [41], [42].

Raman probes are now commercialized by several vendors and applied either on

upstream or downstream cell-based processes. They are currently explored to

Analytics and virus production processes

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