suitable MC choice and should therefore always be investigated in
relation to one another, while Muoio et al. [46] were able to show
that parallel development of MC and chemically defined, serum-
free hMSC expansion medium resulted in enhanced cell attach-
ment, improving the expansion process. Hence, the development
of new chemically defined and serum-free hMSC media will likely
result in the creation of novel MCs specifically tailored to be used in
tandem. To this end, selecting the appropriate MC type for a
particular expansion approach and optimizing the attachment
phase in the bioreactor is as important to the development of the
hMSC cultivation process as the separation of the cells from the
MCs during harvest.
1.3
General
Procedure for and
Results of the Serum-
and Xeno-Free
Expansion of Human
Mesenchymal Stem
Cells from Adipose
Tissue in the BioBLU®
As shown in Fig. 2a, the stem cell expansions were realized using
cells from a working cell bank (WCB), which had been previously
adapted to the culture medium used in the subsequent experi-
ments. Inoculum preparation for all spinner flask (see Subheading
3.2) and bioreactor cultivations (see Subheading 3.3) was per-
formed using T75-Flasks (see Subheading 3.1), which yielded an
average of 8 106 cells per flask with a viability of 97.5% after
3 days, corresponding to an approximate doubling time of 21 h.
Following harvest, the cells of four flasks were pooled to produce
the inoculum. Here, we would like to point out that, with regard to
Working Cell Bank
(WCB)
1
Inoculum production
(in T-flasks)
2
Cell expansions
(Spinner flasks or BioBLU)
3
Cell harvest
4
Cell purification
5
Cell freezing and
storage
6
Screening for the optimum medium-
microcarrier combination
(multiwell plates, spinner flasks)
Spinner flask: Fluid dynamic
characterization and determination of
the suspension criteria, optimization of
cell attachment and harvest efficiency
BioBLU: Fluid dynamic characterization
and determination of the suspension
criteria
(A)
(B)
W
o
r
k
i
n
g
C
e
l
l
B
a
n
(
W
C
B)
1
I
n
o
c
u
l
u
m
(
i
n
T
-
f
l
a
sk
s)
2
C
e
l
l
(
S
p
i
n
3
C
e
ll
4
C
e
l
l
p
u
r
5
C
e
ll
f
r
e
e
z
i
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g
a
n
d
s
t
o
r
a
g
e
6
Fig. 2 A general overview of the steps to be taken for the successful expansion of hMSC cells: which includes
(a) the gradatim approach to the cultivation procedure as described in this book chapter and (b) the
prerequisite experiments, which need to be performed prior to the expansion cultivations
Mesenchymal Stem Cell Expansion at Benchtop-Scale
89