ranging from capacitance measurements to Raman spectroscopy

and enzymatic glucose measurements have become available in

probe format allowing to follow numerous parameters online [20].

2.7

Other

Supplements

pH regulation could be performed by addition of acid or base,

typically HCl and NaOH (see Note 9).

Table 1

Summary table of important parameters

Parameter type

Parameter

Ranges/guidelines (for a 3 L culture)

Physical

parameters

DO percentage

20–50% of dissolved oxygen in the culture (see Subheading

3.4, step 4)

Air injection

In headspace around 100 ccm and through a macrosparger

50 ccm (see Subheading 3.4, step 2)

CO2 injection

5–8% in headspace for equilibration. Could be used in

sparger to regulate the pH (see Subheading 3.4, step 2)

Temperature

37
C could be shifted down (see Subheading 3.4, step 1)

Agitation

Around 200 rpm depending on the coupling system (see

Subheading 3.4, step 3)

pH regulation

Around 6.7–7.5 (see Subheading 3.4, step 5)

Initial volume

Will be based on the final bioreactor volume minus the

volume of feeds and inoculum. Pay attention that this

volume still needs to be above a limit defined by the

supplier mainly to guaranty immersion of probes and the

complete immersion of the impeller to control

generation of foam (see Subheading 3.5, step 1)

pCO2

pCO2 level needs to be carefully monitored especially

during the scale-up. Sensitivity to this parameter will

differ from a cell line to another, usually 40–50 mmHg

[27]

Biological

parameters

Biomass

Initial VCC to be determined regarding process

performances (0.3 to multiple million per mL in case of

perfusion) (see Subheading 3.6, step 2)

Metabolites

concentration/

consumption

Carbon source (e.g., glucose) and by-products (e.g.,

lactate, ammonia) need to be carefully monitored (see

Note 2)

Feeding strategy

Feeds percentage

Depending on the cell line and produced molecule, feeds

percentage could vary. An experiment to determine the

best strategy should be performed in advance. Ranges to

be tested are usually provided by the supplier (see

Subheading 2.3)

Addition strategy

As for the percentage, addition strategy will be based on

the cell’s needs and need to be tested (see Subheading

2.3)

Key components

adjustment

Additional feeding of key metabolites in the culture could

be made on top of feeds addition (see Note 6)

Foam

Foam

Foam level needs to be controlled to avoid clogging of

exhaust filter (see Subheading 3.7, step 4)

6

Ange´ lique Schmid et al.