parallelization of yeast diode and implementation of a concentration gradient
DESCRIPTION
Current Microfluidic designs lack multiple cell growth chambers. Concentration gradients have been explored and developed but there has not been a fusion of multiple cell chambers combined with a concentration gradient. Our objective:To create a microfluidic device with 10 Tesla Diodes, cell growth ports, with an easily controllable concentration gradientTRANSCRIPT
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○ S y s t e m s B i o d y n a m i c s L a b ○
Parallelization of Yeast Diode and Implementation of a Concentration Gradient
BENG 129A: Design Development in Cell Systems Bioengineering
Jeff M. Hasty PH.D.
Group #6: Douglas Cohen, Hirak Desai, Lawrence Hui, Robert Langsner, Rushang Patel
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Microfluidics Applications
Advantages:– reduces time– small requirements of solvents, reagents, and cells– cost effective– versatility in design– parallel operations
Exerts more control over the cellular microenvironment– Chemically– Thermally– Geometrically
Uses:– DNA sequencing and seperation– Enzymatic and Immunoassays– Cell counting and cell sorting– Exploring single cell and its activity– Imaging (cells confined to monolayer)– Montioring gene expression
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Microfluidic Theory
Design principles: Laminar Flow = Linear Design
Re
vLN
4
8 LP Q
r
3
12 LP Q
wd
V I R Ohm’s law
Poiseuille (Laminar) flow
Circular Channel
Rectangular Channel
For water flow at 1mm/s through a channel 100µm wide, NRe = 0.01
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Problem Statement
• Current Microfluidic designs lack multiple cell growth chambers. Concentration gradients have been explored and developed but there has not been a fusion of multiple cell chambers combined with a concentration gradient.
• Our objective:– To create a microfluidic device with 10 Tesla Diodes, cell growth
ports, with an easily controllable concentration gradient
Identify a Need
DesignFlow
AnalysisFinalize Design Fabrication Testing
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2) Draw Up A General Design
• Expansion of the “Tesla microChemostat” design• Monolayer growth in cell chamber
• Creeping media flow in chamber throughout expt• Long runtimes
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LoadRun
Identify a Need Design
FlowAnalysis
Finalize Design Fabrication Testing
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• Microscope Viewing Area:
40mm X 20mm
• Microscope Scope is already
built, thus viewing is limited
to the restraints of the microscope.
Standards and Constraints
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• Design Restraints– Flow Rates must match at each node to ensure a 50%-50%
mixture.– Lengths must be sufficiently long to allow complete mixture
of media by diffusion. (Dependant on flow rates)– Inputs pressurized by gravity is limited in range (5 – 40
inh20)– Inputs pressurized by Air Pumps have much higher ranges
but limited to two distinct levels (ie. 100 and 150 inh20)– Chip can only sustain 15 PSI (normal pressures are 0.6 – 1
PSI)
Standards and Constraints
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Standards and Constraints (Contd.)
Linear Media Concentration Gradient
1 2 3 4 5 6 7 8 910
20
30
40
50
60
70
80
90
100
chamber #
Con
cent
ratio
n %
Tree Design
1 2 3 4 5 6 7 8 90
10
20
30
40
50
60
70
80
90
100
chamber #
Con
cent
ratio
n %
Serial Design w/ 9 channels
1 1.5 2 2.5 3 3.5 4 4.5 510
20
30
40
50
60
70
80
90
100
chamber #
Con
cent
ratio
n %
Serial Design w/ 5 channels
Media Mixing Efficiency
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Parallel: The parallel chip design is modeled after a parallel electric circuit. Originally used for a cell culture array for mammalian cells, we tried to expand on this design to include a chemical gradient. Each cell chamber is independent from the rest, therefore it increases the ease of fabrication and analysis of flow rates.
Serial Dilution: This design capitalizes on constant media/buffer dilution to create discrete concentration levels. The media is diluted after each successive chamber. This design decreases the amount of work to analyze and model the circuit significantly. However, with this design the gradient is no longer linear, rather it becomes logarithmic as the M (media) mixes with B (buffer) at each successive port.
Tree Dilution: This design allows us to create linear gradient with 3 media inputs and 5 ports overall. The was a novel design drawn by Group 6.
Alternative Designs
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Parallel
Slant Resistors
Fluid Rectifiers
Serial
Tree Dilution
Square Resistors
Slant Resistors
Fluid Rectifiers
Square Resistors
Serial Dilution
Basic Concept
Design Thought Process
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Features
Parameters Weight
Parallel
Design
Tree Design w/ slant
resistors
Tree Dilution
w/ fluid rectifiers
Serial Dilution w/ square resistors
Cell Loading Complications 10 1 3 3 3
Media mixing efficiency 10 4 3 2 4
Flow Analysis 8 4 0 0 3
# of inputs 7 4 3 3 4
Ease of Fabrication 6 4 4 4 4
Spatial Flexibility 6 3 2 2 4
Media gradient efficiency 5 4 4 4 4
Modification Aptitude 4 4 2 2 3
Cost 4 4 4 4 4
Ease of Design 3 3 3 3 3
Industrial Application 2 2 2 2 2
Total 217 174 164 2310 = inadequate; 1 = weak; 2 = sufficient; 3 = good; 4 = excellent
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3) Flow Analysis for General Design
MATLAB scripts facilitate chip design process by calculating pressures/flows throughout device
Identify a Need Design
FlowAnalysis
Finalize Design Fabrication Testing
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Matlab: MOCAMicrofluidic Open Circuit Analyzer
Our system is modeled to the right, with hypothetical inputs (power sources) and arbitrary segment lengths (resistances). With 4 unknown inputs and 35 segment lengths, it becomes nearly impossible to solve this system by hand.
On the right is an example of a circuit in which the currents at each joining section do not match and the power inputs are off so that a back-flow occurs on the far right.
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Flow Analysis (correct)
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Final Design
Linear Media Concentration Gradient
1 2 3 4 5 6 7 8 910
20
30
40
50
60
70
80
90
100
chamber #
Con
cent
ratio
n %
Tree Design
1 2 3 4 5 6 7 8 90
10
20
30
40
50
60
70
80
90
100
chamber #
Con
cent
ratio
n %
Serial Design w/ 9 channels
1 1.5 2 2.5 3 3.5 4 4.5 510
20
30
40
50
60
70
80
90
100
chamber #
Con
cent
ratio
n %
Serial Design w/ 5 channels
Media Mixing Efficiency
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Final DesignFinal Design: Cell LoadingFinal Design: Media Running
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Mask glued to
borosilicate glassMask file to be
sent for printing
Process Timeline: Next Quarter
Identify a Need Design
FlowAnalysis
Finalize Design Fabrication Testing
Fabrication (3 Step Process)
1. Print Photomasks
2. Fabricate Mold
3. Produce Chips from Mold
Patterned Silicon Wafer Finished Devices!
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Microfluidic device fabrication can be broken down into 3 steps:
Microfluidic Overview
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Process Timeline: Next Quarter
Identify a Need Design
FlowAnalysis
Finalize Design Fabrication Testing
S. Cerevisiae
strain K699
E. coli wt
strain JM2.300
Automated Microscope
Testing (Dye => Media => Cells)
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Acknowledgements
PI
Jeff M. Hasty PH.D.
Graduate Students
Lee Pang
Scott Cookson
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