bioreactor design and implementation for generating
TRANSCRIPT
Bioreactor Design and Implementation for Generating
Anterior Cruciate Ligament Bioscaffolds Capable of Supporting Chondrogenic Cell Differentiation
Megan Barnum Sarah Dwyer Lauren Hoft Melinda Hunter Lacey Prestwood
Dr. Mandi Lopez Department of Veterinary Clinical Science Dr. Daniel Hayes Department of Biological and Agricultural Engineering
Problem Statement
Background Information
Significance of Bioscaffold
Overall System Layout
Product Design Specifications
Chamber Design
Oxygen Reservoir Design
Additional Parts
Materials & Cost
Measurable Objectives
Constraints
Design Tools Pugh Selection Chart Physical Decomposition Chart House of Quality Gantt and PERT charts
Future Work
To design a bioreactor capable of loading adult derived adipose stem cells onto three-dimensional scaffold structures while holding tension and maintaining a
sterile environment.
Bioreactor Function Synthesize bioscaffold
Stem cells Media
Maintain sterile environment Aseptic media exchange No leakage at connections
Maintain sufficient oxygen supply
Previous bioreactor design Inability to maintain sterile
environment System leaks at connection
points No internal cell growth
Synthetic
Highly porous
Extracellular matrices
Facilitates growth and regeneration of mammalian cells
Scaffold Structure
Bird’s eye view of scaffold 10 mm x 30 mm
Commonly Practiced Procedure
Skin graft Remove skin fascia from
lower leg
Construct into cylinder shape
Exchange with torn ACL through orthoscopic surgery
Synthetic Bioscaffold Exchange with torn ACL
through orthoscopic surgery
Higher success rate Designed with patient in
mind
Improved Procedure via Bioscaffold
Overall System Layout
Pump
Filter Filter
Oxygen Reservoir
Chamber
Stopcock Stopcock
Chamber
Oxygen Pump
Laptop with fluid flow calculation software
*arrows denote tubing
Environment Steady state
Fluid flow Oxygen exchange Gas pressure Temperature
Sterile
Chamber Cylindrical Airtight 50 mm x 20 mm 45° twist
Filtration Thermo scientific:
Hydrophobic Filter
Oxygen Reservoir
Elliptical
45.85 mm x 20.45 mm x 7.75 mm
Tubing: Tygon R3603
7.0 mm diameter
Remove and Replenish Nutrients
Oxygen reservoir
Aseptic
Every three days
Overall System Layout
Pump
Filter Filter
Oxygen Reservoir
Chamber
Stopcock Stopcock
Chamber
Oxygen Pump
Laptop with fluid flow calculation software
*arrows denote tubing
Mechanical aspects of design 45° twist Tension kept on
scaffold
Ways to achieve important elements 45° twist – plastic
screw on caps Tension – tension
rods, gear locking mechanism
Chamber design with tension
rods and gasket
Initial Design
Front view
Expanded view
• Adapted to the system with luer locks and stopcock.
• The top is connected to a stopcock
• Bottom is connected to a barbed luer lock which is connected to tubing.
Overall System Layout
Pump
Filter Filter
Oxygen Reservoir
Chamber
Stopcock Stopcock
Chamber
Oxygen Pump
Laptop with fluid flow calculation software
*arrows denote tubing
Top
Bottom
Full Assembly with filter flap and stir bar
Promotes gas exchange and replenishes oxygen throughout medium.
Shape prevents stagnation points and cell losses.
Constructed using ABS Plastic
Stir bar (12.7mm x 3.2mm) to agitate fluid and gas.
Filter to release negative pressure and gases Prevent negative pressure
Promote waste gas removal
Volume = 30,430.5 mm3 = 30.43 ml ◦ Will hold 5-10ml of media needed to pass through system.
Stir bar (12.7mm x 3.2mm) ◦ Octagonal with pivot ring to
maintain stability while inside OR
Barbed luer locks: male and female ◦ Used to attach Prototype I to
medical grade tubing
◦ Composed of Nylon
◦ Sealed to Prototype I using Silicone sealant to prevent leakage
Pump: Uses air to force fluid back
and forth through scaffolding and oxygen reservoir
Stopcock: Allows fluid to be added and removed
from the system aseptically
Air filter: Filters air entering the system from
pump to maintain sterility in tubing
Overall System Layout
Leakage points
ITEM COST Budget
Nylon Female Luer Lock to Barb Connector (5/32 inch)
$9/pack of 25 1 pack $9
Tygon R3603 Tubing (5/32 inch)
$0.39/ft 12 feet $4.68
3-D print oxygen reservoir prototypes
$200 each 3 prototypes $600
Stir bar rods $4 each 3 stir bar rods $12
Chamber (3/4 inch clear PVC) $2.13/ft 1 foot $2.13
Rubber stoppers $0.60 each 4 stoppers $2.40
Tension rods (stainless steel rod) $12/3 ft 1 rod $12
Cells $400/million 2 million $800
Filters free available in lab
Stopcocks free available in lab
Air pump free available in lab
Oxygen pump free available in lab
Media free available in lab
TOTAL: $1,442.21
Cell adhesion Factors
External and internal cell proliferation
Plan to execute Scanning electron microscope images of
structure
Flow rate Factors
Constant perfusion with varying levels of pressure
Maintain airtight seal throughout system
Plan to execute Use fluid flow calculation software Monitor and note possible leakage.
Capacity of pressure release valve for oxygen reservoir Factors
Flow rate of fluid Volume of reservoir Amount of oxygen supplied to reservoir Speed of agitation by stir bar
Plan to execute Test oxygen supply against negative
pressure Pressure sensor
Tension required for scaffold
Factors
Density and flow rate of fluid
Scaffold attachment to tension rods
Gear locking mechanism
Withstand 70 N of force
Plan to execute
Measure density of fluid
Calculate flow rate
Trial and error testing various tension settings
Budget $2050
Time
Team collaboration (group of 5)
Materials Finding available materials to fit design requirements
Pugh Selection Chart
Physical Decomposition Chart
House of Quality
Gantt and PERT charts
A. Chamber with added tension rods E. OR with flat bottom, vent component, aeration pump
B. Chamber with rubber stoppers on ends to stop leakage F. OR with pressure relief valve, rubber stoppers, aeration pump
directly through medium
C. Chamber with external threaded attachments to allow for
tubing and hardware connections.
G. OR with holes approximated for standard luer locks (tubing
fittings); syringe attachment with flap, aeration at top of device
D. Chamber with two threaded elements to allow for the 45˚ twist
in scaffold to be created.
H. OR with external threaded attachments for tubing connections,
pressure relief flap, aeration pump at bottom through the medium
Row Criteria Concept
A B C D E F G H
1 COST
D
A
T
U
M
+ + +
D
A
T
U
M
+ + -
2 FUNCTIONALITY + + + - + +
3 SIMPLICITY OF DESIGN - + - + + -
4 AVAILABILITY OF MATERIALS + - + - - +
5 EASE OF MANUFACTURING - + + + + -
6 EASE OF ASSEMBLY - + + + + +
7 ABILITY TO PROTOTYPE - + + - + -
8 TENDENCY TO LEAK - - - - + +
9 AESTHETICS + + - - + +
PLUSES 0 4 7 6 0 4 8 5
MINUSES 0 5 2 3 0 5 1 4
Bioreactor
Chamber
Filters Tubing
Oxygen Reservoir
Gas Exchange pump
Power Pump
Fluid Transport
Open/Close Switch
Tension Rods
House of Quality Engineering Characteristics
Improvement Direction const const const
Units n/a n/a in3/s lb/N mm
Customer Requirements
Imp
ort
ance
Wei
gh
t F
acto
r
Ste
rili
ty
Co
nst
ant
Pro
fusi
on
of
Med
ian
Aer
atio
n
Ten
sio
n
Sh
ape
of
cham
ber
Par
t re
liab
ilit
y
Cost 5 9 9 3 1 1 9
Ease of Use 4 3 0 3 9 0 9
Functional 5 9 9 9 9 9 9
Power Consumption 3 0 3 1 0 0 3
Maintenance 5 9 3 1 3 1 9
Durability-life span 4 9 1 3 9 1 9
Raw Score 192 118 92 137 59 216
Relative Weight % 23.59 14.5 11.3 16.83 7.25 26.53
Rank Order 2 4 5 3 6 1
House of Quality Engineering Characteristics
Improvement Direction const Const const
Units n/a n/a in3/s lb/N mm
Customer Requirements
Imp
ort
ance
Wei
gh
t F
acto
r
Ste
rili
ty
Co
nst
ant
Pro
fusi
on
of
Med
ian
Aer
atio
n
Ten
sio
n
Sh
ape
of
cham
ber
Par
t re
liab
ilit
y
++ Strong positive
+ Positive
None
- Negative
-- Strong Negative
++
+
+
- +
-
Manufacture Prototype I
Create efficient system
Test Prototype I Design
Chamber
Airtight
Aseptic
Ability to culture scaffold
Oxygen Reservoir
Proper gas exchange
Reduce material of structure to lower production cost
Add pressure relief valve if needed
Grow viable bioscaffold
Questions?