SNF Grand RoundsJuly 13, 2006
ME342
Jennifer Blundo
Gretchen Chua
Yong-Lae Park
Ali Rastegar
Project Goal
• Design a bioMEMs substrate to apply and
measure electromechanical forces in the
differentiation of human embryonic stem cell-
derived (hESC)-cardiac myocytes (CM)
Undifferentiated hESCs-Fluc-eGFP
(DAPI nuclear stain)
hESC-CMs organized in embryoid body
bioMEMS device
Contractility
Electrophysiology
Mechanical force
Current Microscale Devices
Thin-film stretchable (0—15%) gold electrodes (25nm) on PDMS. Lacour et al, 2005.
Thin-film gold strain gauges (200nm) encapsulated in PDMS (50μm). Wen et al, 2005.
64 Electrode array for extracellular recording, Multi Channel Systems
Pressure actuated PDMS membrane (120μm) with S-shaped SiO2 traces. Lee et al, 2004.
BioMEMS: Engineering Specs
Device Requirement Target Value
1. Apply mechanical strain Up to 10%
2. Apply electric field ~O(1) V/cm
3. Measure electric potential (ECG) 100μV—1mV
4. Area of mechanical deformation A < 1cm2
5. Size of electrodes diameter = 20μm
6. Inter-electrode spacing spacing = 250μm
7. Area of cell culture A > 1cm2
8. Thickness of substrate t < 1mm
BioMEMS: Device Design
A. Unstrained state B. Strained state
Glass/Quartz: Optically transparent baseplate PDMS: A biocompatible elastomeric polymer
PPS: A biocompatible elastomeric polymer
Ti: Adhesion layer for electrodes
Gold: Biocompatible thin film electrodes
SU-8: Transparent polymer
Mechanical Strain—In Vitro Model
• Goal: To apply cyclic mechanical strain to hESC precursor cells
and observe differentiation
BioMEMS: Stretchable Electrodes
C. S. Park, M. Maghribi Characterizing the Material Properties of Polymer-Based Microelectrode Arrays for Retinal Prosthesis
Biaxial Loading—10% Strain
• Material Properties– PDMS: E = 500kPa, v = 0.5
– Gold: E = 78GPa, v = 0.44
Stress Contour Plot Strain Contour Plot
• Geometry– PDMS: t = 100μm
– Gold: t = 100nm, w = 30μm, L = 240μm, pitch (p) = 120μm
PDMS & Gold Electrode Strain
BioMEMS: Loading Curves
• Operating pressure < 15psi
• Young’s Modulus PDMS E = 500kPa
• Thickness = 100um
• Membrane diameter = 1cm
• Loading post diameter = 0.7cm
Elongation vs. Modulus of PDMS Membrane (100um thickness)
0
5
10
15
20
25
30
35
0 5 10 15 20 25
Applied Pressure (psi)
Elo
nga
tion
(%
)
250 kPa
500 kPa
1500 kPa
3000 kPa
Fabrication: Baseplate
Step 1: Clean Pyrex 7740 4” glass wafer (300μm thick), dehydrate
5min @ 200°C
Equipment: Acetone/Methanol/IPA/DI rinse Location: MERL
Glass
Fabrication: SU-8 Processing
Glass
Channels to apply vacuum pressure to PDMS membraneGlass
Exposed SU-8
Unexposed SU-8
Step 2: Spin 1st layer SU-8-100 (100μm thick), prebake 10min @
65°C, softbake 30min @ 95°C, expose, postbake 1min @ 65°C,
10 min @ 95°C
Equipment: Spin coater, hot plate, exposer Location: MERL
Fabrication: SU-8 Processing
Step 3: Spin 2nd layer SU-8 (100μm thick), prebake, expose,
postbake
Equipment: Spin coater, hot plate, exposer Location: MERL
Loading post to support PDMS membrane
Glass
Exposed SU-8
Unexposed SU-8
Fabrication: SU-8 Processing
Step 4: Spin 3rd layer SU-8 (100μm thick), prebake, expose,
postbake
Equipment: Spin coater, hot plate, exposer Location: MERL
Glass
Exposed SU-8
Unexposed SU-8
Fabrication: SU-8 Processing
Step 5: Spin 4th layer SU-8 (80μm thick), prebake, expose,
postbake
Equipment: Spin coater, hot plate, exposer Location: MERL
Glass
Exposed SU-8
Unexposed SU-8
Fabrication: SU-8 Processing
Step 6: Develop SU-8, IPA/DI rinse
Equipment: Location: MERL
Glass
Exposed SU-8
Fabrication: SU-8 Processing
Step 7: Pipette tetrafluoropolymer (PS200 or T2494) to prevent
PDMS membrane stiction
Equipment: Location: MERL
Glass
Exposed SU-8
Tetrafluoropolymer
Fabrication: Baseplate Assembly
Step 8: Laser cut Pyrex 7740 4” quartz wafer (300μm thick) and
bond quartz over SU-8
Equipment: Laser cutter Location: MERL
Glass/Quartz
Exposed SU-8
20μm clearance between loading post and PDMS membrane
Fabrication: PDMS Membrane
Step 1: Clean 4” silicon wafers
Equipment: wbnonmetal Location: SNF
Silicon
Fabrication: PDMS Membrane
Step 2: Spin sacrificial layer 5% (w/v) poly(acrylic acid) (PAA)
(3000 rpm, 15 s) and bake (150C, 2 min)
Equipment: Spin coater, Hot plate Location: MERL
Silicon
PAA
• Advantages of water-soluble films – Deposited by spin-coating– The solvent removed at a low temperature (95–150C)– The resulting layer can be dissolved in water– No corrosive reagents or organic solvents– Faster release of features by lift-off
• Compatible with a number of fragile materials, such as organic polymers, metal oxides and metals—materials that might be damaged during typical surface micromachining processes
Sacrificial Layers—PDMS Micromachining
Sacrificial Layers—PAA & Dextran
Fabrication: PDMS Membrane
Step 3: Spin 20:1 Sylgard 184 poly(dimethylsiloxane) (PDMS)
(40μm thick), bake (60C, 1 hr), O2 plasma
Equipment: Location: MERL
Silicon
PAA
PDMS
2mm gap at edge of wafer to prevent lift-off of PDMS during processing
Fabrication: Electrode Array
Step 4: Align beryllium copper shadow mask and temporarily
bond.
Equipment: EV aligner Location: SNF
Silicon
PAA
PDMS
Shadow Mask
Ti
Au
20μm diameter electrodes
30μm width tracks for electrode connections
Fabrication: Electrode Array
Step 5: Evaporate Ti adhesion layer (10nm thick) and Au layer
(100nm thick)
Equipment: Innotec Location: SNF
Silicon
PAA
PDMS
Shadow Mask
Ti
Au
20μm diameter electrodes
30μm width tracks for electrode connections
Fabrication: Electrode Array
Step 6: Remove shadow mask, O2 plasma
Equipment: Drytek Location: SNF
Silicon
PAA
PDMS
Shadow Mask
Ti
Au
Fabrication: Electrode Array
Step 7: Prebake 110°C, spin photo-patternable silicone (PPS)
WL5153 30sec @ 2500rpm (6μm thick), expose*, postbake @
150°C**, develop
Equipment: Hot plate, Spin coater, Karl Suss*, BlueM oven**,
wbgeneral
Silicon
PAA
PDMS
Shadow Mask
Ti
Au *Proximity exposure
**Need to characterize in BlueM Oven
PPS
Fabrication: Electrode Array
Step 8: Dissolve sacrificial layer PAA in water
Equipment: wbgeneral Location: SNF
Silicon
PAA
PDMS
Shadow Mask
Ti
Au
PPS
Fabrication: Electrode Array
Step 9: Air dry device
Equipment: N2 gun
Silicon
PAA
PDMS
Shadow Mask
Ti
Au
PPS
Fabrication: Assembly
Step 1: O2 plasma PDMS and quartz surfaces
Equipment: Drytek
Silicon
PDMS
PPS
Ti
Au
Glass/Quartz
SU-8
Fabrication: Assembly
Step 2: Bond PDMS membrane to glass
Glass/Quartz PDMS
PPS
Ti
Au
SU-8
Next Steps
• Transparency masks SU-8 molding
• Laser cutting quartz
• Plate electrodes on PDMS
• Finish SNF training
Acknowledgements
Sacrificial Layers—PDMS Micromachining
• Challenge: etchants may diffuse through PDMS membrane—
these traces may ultimately by harmful to cell culture
• Photoresist—acetone removal through selectively
etched holes
• Backside etch stop of 4000A thick SiO2—1 mm
thick PDMS membrane coated with 540A thick
sputtered Cr covers PDMS membrane and a
PDMS mold is created to protect the whole
PDMS structure.
• Water soluble sacrificial layers—dextran and
PAA—insoluble in most organic solvents!
Sacrificial Layers—PDMS Micromachining
Sacrificial Layers—PAA & Dextran
•Films were prepared by spin-coating (3000 rpm, 15
s) from a 5% (w/v) polymer solution in water.
•Films were then dried by placing the substrates on
a hot plate at 150C for 2 min.
Stimulation Electrodes
• Goal: To pattern gold electrodes within a flow
chamber for selectively stimulating hESCs– Electrodes 100μm x 5000μm (10 per well)– Interelectrode distance 1000μm– Contacts pads 2mm x 2mm (10 per well)
• Polished glass wafers 1 mm thick
BioMEMS: Strain gauge
• Need a strain gauge and a reference strain
gauge for every deformable area.
Strain gauge design
• Length (L)
• Trace width (w)
• Number of turns (t)
• Distance between turns (p)