between various bioreactor scales. For the BioBLU^® 0.3c, these

ratios are 0.5, 1.2, and 1.8 for d/D, HL/D, and H/D, respectively.

More information regarding the dimensions of the various com-

mercially available BioBLU^® bioreactor types may be found on the

manufacturer’s website. Furthermore, due to the low stirrer instal-

lation height, high relative velocities are obtained near the bottom

of the reactor, which allows the suspension of MCs at low power

inputs and thus at acceptable hydrodynamic loads for the cells.

As previously mentioned, allogenic therapies require large

quantities of cells to treat multiple patients using a single universal

donor. This necessitates the use of pilot or production scale stirred

single-use bioreactors to achieve therapeutically relevant cell den-

sities in an economic and ecological manner. Due to the limitations

faced by spheroid-based cultures and the associated lack of process

reproducibility, this technique is of diminished importance here. In

contrast, the improved reproducibility when cultivating with MCs,

means their application, especially with regard to the production of

hMSCs for subsequent use in allogeneic therapies, is very popular.

Nevertheless, the same aspects (e.g., cell attachment, medium

composition), as described for the benchtop systems, apply at this

scale also. In their studies, Schirmaier et al. [37], Jossen et al.

[6, 38], and Lawson et al. [39] achieved maximum cell densities

of up to 0.7  10⁶ cells mL^1 when cultivating in stirred pilot-scale

bioreactors (Biostat STR^® 50 L and Mobius^® CellReady 50 L) and

using cell culture medium supplemented with 10% human platelet

lysate and 5% fetal bovine serum (FBS), respectively. While it has

been shown that maximum cell densities (0.04–0.4  10⁶ cells

mL^1) and expansion factors achieved in stirred bioreactors using

xeno- and serum-free cell culture media are still lower than those

achieved in serum-containing medium, a wide variety of new

serum-free media have recently come to market, paving the way

for higher hMSC densities to be achieved on production scale in

future.

1.2

Chemically

Defined, Serum-Free

Expansion of hMSCs

The most conventional cell culture media used for the production

of hMSCs are based on defined basal media, such as Dulbecco’s

Modified Eagle Medium (DMEM), Roswell Park Memorial Insti-

tute (RPMI) 1640, or Minimum Essential Medium (αMEM).

These basal media are subsequently supplemented with expensive

additives such as (a) proteins for cell adhesion, (b) lipids for cellular

anabolism, (c) growth factors and hormones to promote cell

growth, and in most cases, (d) an additional 10–20% (vol/vol)

FBS. While the disadvantages of serum addition (e.g., high batch-

to-batch variability, possible contamination with prions, viruses, or

zoonotic agents [40–44]) have been known for years, serum-

containing media are still used in many published academic studies

to this day. Meanwhile chemically defined, serum-free media not

only offer the advantage that all their components are known but

also that their quantities are stoichiometrically defined, facilitating

Mesenchymal Stem Cell Expansion at Benchtop-Scale

87