16
Downstream
thirty seven
bioMérieux Boxtel, The Netherlands, has successfully automated a process for MAb purification
by running a STREAMLINETM expanded bed adsorption column on an ÄKTApilotTM system.
This new approach has reduced their production times and saved on investments.
Two-step automated MAb
purification on STREAMLINE
and ÄKTApilot
An interview with Ruud van Meersbergen
and Bert Timmers from bioMérieux Boxtel, The Netherlands.
T
he advantages offered by STREAMLINE expanded
bed adsorption (EBA) and the new sanitary
ÄKTApilot system controlled by UNICORNTM are
benefiting a company in The Netherlands with their
production times for a monoclonal antibody used in
diagnostics. DownstreamTM recently talked with Ruud van
Meersbergen and Bert Timmers to learn about their process
and the benefits gained.
Their process begins with fermentation. Hybridoma cells are
grown in a bioreactor (equipped with acoustic resonance) in
serum-free culture medium. Harvest is
performed by continuous perfusion. The
harvest is pre-filtered and conditioned
before purification via an automated
two-step procedure using STREAMLINE
rProtein A and in-line desalting on
SephadexTM G25 Medium. Both columns
are connected to an ÄKTApilot system.
The final product is filtered through a
0.2 µm filter.
Why are you running your
process on ÄKTApilot and
STREAMLINE?
"To fulfil our needs (i.e. yearly production
of 400 g of MAb), we had to change our
culturing process from a hollow-fiber
system to a bioreactor. Because of the
lower expression level in the bioreactor, we have to handle
large volumes of harvest. Centrifugation and filtration is not
an option for us with these large volumes. And that is the
reason we chose EBA. Time-consuming centrifugation and
filtration of harvest were not needed, and expensive investment
in such equipment was avoided. Additionally, the capacity of
the ÄKTApilot system matche our intended production scale,
and since ÄKTApilot seemed to be compatible with
STREAMLINE columns, it was an obvious choice for us."
Can you describe the first steps?
"The volume of the bioreactor is 15 liters and we harvest
about 25 liters/ day. Cell density is about 1-2x10
7
.
Approximately 8 days of harvest are pooled, pre-filtered and
then loaded on the STREAMLINE 100
column. The column contains 1.3 liters
of STREAMLINE rProtein A adsorbent,
which is equivalent to a sedimented bed
height of 17 cm.
For the EBA step: equilibration and
sample loading are run at approximately
220 cm/h, elution is performed in
packed bed mode at 100 cm/h. The
binding capacity of the column is about
15 mg/ml and recovery is minimal - 80%.
The desalting column (INdEXTM 200)
has a volume of 11 liters. Sample is
loaded at 100 cm/h with elution at
70 cm/h. The target molecule is recovered
in about 2 liters.
The volumes of both columns are set in
such way that the Sephadex G-25 Medium is never loaded
with more than 15% of the bed volume of that column,
which results in a resolution factor > 1.25 between the
conductivity- and A
280
-peaks."
ÄKTApilot system has been configured to run
STREAMLINE 100 column and INdEX 200 in an
automated MAb process.
