snf grand rounds july 13, 2006 me342 jennifer blundo gretchen chua yong-lae park ali rastegar

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


Step 3: Spin 2nd layer SU-8 (100μm thick), prebake, expose,

postbake


Loading post to support PDMS membrane

Glass

Exposed SU-8

Unexposed SU-8


Step 4: Spin 3rd layer SU-8 (100μm thick), prebake, expose,

postbake


Glass

Exposed SU-8

Unexposed SU-8


Step 5: Spin 4th layer SU-8 (80μm thick), prebake, expose,

postbake


Glass

Exposed SU-8

Unexposed SU-8


Step 6: Develop SU-8, IPA/DI rinse

Equipment: Location: MERL

Glass

Exposed SU-8


Step 7: Pipette tetrafluoropolymer (PS200 or T2494) to prevent

PDMS membrane stiction


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


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


Step 3: Spin 20:1 Sylgard 184 poly(dimethylsiloxane) (PDMS)

(40μm thick), bake (60C, 1 hr), O2 plasma


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


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


Step 6: Remove shadow mask, O2 plasma

Equipment: Drytek Location: SNF

Silicon

PAA

PDMS

Shadow Mask

Ti

Au


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


Step 8: Dissolve sacrificial layer PAA in water

Equipment: wbgeneral Location: SNF

Silicon

PAA

PDMS

Shadow Mask

Ti

Au

PPS


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


• 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—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)

snf grand rounds july 13, 2006 me342 jennifer blundo gretchen chua yong-lae park ali rastegar

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