Automating the Cell Culture
Sampling Process
Mike Phipps
Tara Ryan
BME 273
March 18, 2002
Problem/Background
• Cell cultures maintained in bioreactors for Research and
Development purposes in pharmaceutical companies must
be sampled regularly
• Samples (10-15mL) are typically taken once most days,
and twice every three days or so when the culture is split
• methods of manually withdrawing a sample from the
bioreactor can be reliable but still come with risks of
culture contamination
• lab workers must be trained and experienced in sterile
technique
Existing Sampling Methods
DO sparger
pH probe
Temperature
probe
Agitator
Sampling syringe
ethanol
Hot plate to maintain temperature
Existing Sampling Methods
DO sparger
Temperature probe
agitator
pH probe
Water out
Water in
Sampling port
Sampling syringe
3-way valve
ethanol
Water gasket for temperature control
Flowchart of the Sampling
Process
Ensure
sterility of
syringe tip
Obtain new
syringe
Make sure tip is
okay to enter
culture
Draw sample from
culture
Insert
syringe tip
into culture
Dispose of
used
syringe
Move collecting
tube to analysis
machines
Pull sample
into syringe
tube
Deposit sample
into collecting
tube
Move syringe
to collecting
tube
Remove
syringe tip
from culture
Project Goals
• reduce the risk of contamination that
occurs due to sampling
• reduce the time it takes a lab worker
to draw a sample from a culture
• reduce the skill and training required
by a lab worker
Design Ideas
Idea #1
• Continuous flow of
medium and cells through
tubing loop
• switch 3-way valve to the
sampling line in order to
draw a sample
• simple
• does not avoid the
traditional syringe switch
Assessment of Design Idea #1
Advantages:
• simple
• inexpensive
• easy setup
Disadvantages:
• does not avoid the
“syringe switch”
• does not reduce the time
or labor needed to sample
Design Ideas
Idea #2
• Ethanol and wash sterilize the syringe tip (needle)
• Use of septa
• Expand to a set of 4 bioreactors
Assessment of Design Idea #2
Advantages:
• Very little risk of
contamination
• Can enclose/sample many
bioreactors
• Reduces the labor/time
needed to sample
Disadvantages:
• Ethanol and wash supplies
must be changed
frequently
• Expensive
• Chance of alcohol residue
on syringe tip (can kill
cells and influence
viability counts)
Design Ideas
Idea #3
• Open flame sterilizes the syringe tip (needle)
• Use of septa
• Water-gasket bioreactor system for better maintenance of the
culture’s temperature
• Expand to a set of 4 bioreactors
Assessment of Design Idea #3
Advantages:
• Very little risk of contamination
• Can enclose/sample many
bioreactors
• Reduces the labor/time needed
to sample
• Once cooled, syringe tip is safe
to enter the culture (you can
calculate how long the tip needs
to cool off after submergence in
the flame, but in #2, there is no
easy way of making sure all the
alcohol wash is gone)
Disadvantages:
• Expensive
• Heat from flame may influence
temperature of hood
environment or of culture
• No flammable
materials/chemicals should be
in the hood
Design Ideas
Idea #4
• Simpler (fewer steps for mechanical arm)
• Reliance on hood to provide sterility
• Expand to a set of 4 bioreactors
Assessment of Design Idea #4
Advantages:
• Reduces the risk of
contamination
• Can enclose/sample many
bioreactors
• Reduce the labor/time
needed to sample
• The hood air source only
blowing when the door
flap is open
Disadvantages:
• Expensive
• Reservoir of new syringes
is briefly exposed to
outside environment each
time a sample is
transferred to a collecting
tube
Conveyor Belt Idea
• Conveyor belt would transport multiple bioreactors
to a stationary mechanical arm so that arm will not
require a track along which it can move
• Cost of a 12-feet long conveyor belt with a
diameter/width of 30 inches is estimated to be
$6000*
• Air inlets (nitrogen, oxygen, etc.) come from pipes
running down from the ceiling; can’t easily move
these with the bioreactor
* according to http://www.matche.com/EquipCost/Conveyor.htm
Final Design
• Combination of Design Ideas #3 and #4 (uses flame
sterilization with the movable door feature)
Advantages:
• Reduces the risk of contamination
•Can enclose/sample many bioreactors
•Reduce the labor/time needed to sample
Disadvantages:
• Heat from flame may influence
temperature of hood environment or
of culture
•The hood air source only blowing when
the door flap is open
• No flammable materials/chemicals
should be in the hood
•Once cooled, syringe tip is safe to enter
the culture (you can calculate how long the
tip needs to cool off after submergence in
the flame, but in #2, there is no easy way of
making sure all the alcohol wash is gone)
• Reservoir of new syringes is briefly
exposed to outside environment each
time a sample is transferred to a
collecting tube
Current Work
• Components of the Device:
– test tubes, test tube rack, disposable syringes, syringe
disposal bag, bioreactor system(s) with septa embedded
in top, hood device with movable divider, mechanical
arm, track, air purifier, Bunsen burner, natural gas line
• Determine the manufacturers, material
composition, price, dimensions, and weight of
each component
Future Work
• Calculations (heat transfers, air
flows, etc.) and research of parts
manufacturers to determine
specifications of the final design
• Draw final design using
AutoCAD
• Production of a prototype?
References
• ABEC Website, <http://www.abec.com>
• B. Braun Biotech Website, <http://www.bbraunbiotech.com>
• Bailey, James E., and Ollis, David F. Biochemical Engineering
Fundamentals. McGraw-Hill Inc.: St. Louis, 1986.
• Balcarcel, R. Robert. Associate Professor of Chemical Engineering,
Vanderbilt University.
• New Brunswick Scientific Website, <http://www.nbsc.com>
• Todar, Kenneth. “The Control of Microbial Growth.” 21 September
2000
<http://www.bact.wisc.edu/microtextbook/ControlGrowth/sterilization.
html>