ME342 Jennifer Blundo Gretchen Chua Yong-Lae Park Ali Rastegar SNF Grand Rounds July 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) hESC-CMs organized in embryoid body Contractility Electrophysiology Mechanical force Undifferentiated hESCs-Fluc-eGFP (DAPI nuclear stain) bioMEMS device

Current Microscale Devices Thin-film gold strain gauges (200nm) encapsulated in PDMS (50μm). Wen et al, 2005. Thin-film stretchable (0—15%) gold electrodes (25nm) on PDMS. Lacour 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 Geometry PDMS: t = 100μm Gold: t = 100nm, w = 30μm, L = 240μm, pitch (p) = 120μm Material Properties PDMS: E = 500kPa, v = 0.5 Gold: E = 78GPa, v = 0.44 Stress Contour Plot Strain Contour Plot

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

Fabrication: Baseplate Step 1: Clean Pyrex 7740 4” glass wafer (300μm thick), dehydrate 5min @ 200°C Equipment: Acetone/Methanol/IPA/DI rinse Location: MERL Handle wafer is Au-coated Si wafer 1. Si wafer Sacrifical layer—ie, thin film of Ti or Au. The metal firm serves as a seed layer for electroplating and facilitates removal of the pdms membrane from the wafer 2. *note need a 2 mm gap at edge of wafer to prevent lift-off of pdms layer during processing 3. Spin thick photoresist (30um) 4. Expose so that elecrodes are raised features 5. Spin PDMS at spin rate to keep membrane thickness below pattern height 6. cure pdms Clean off any pdms adhered to electrodes 7. Electoplate metal electrodes, strain gauges, and contact pads-Pt or Au 8. O2 plasma-oxidize PDMS surface, allows PR to wet the PDMS surface, eliminating beading and ensuring a smooth, uniform coat of resist. 9. Spin resist 10. Expose area for traces. 11. Re-plasma 12. ITO adhesion layer (20nm) 13. E-beam evaporate gold film (150nm) 14. PR removal-acetone lift-off 15. Ethanol rinse and dry to prepare for passivation 16. O2 plasma 17. Spin PDMS membrane 18. Cure PDMS on top of glass substrate Glass

Fabrication: SU-8 Processing 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 SU-8 is a negative resist, exposed areas stay (see Signs) 1. Si wafer Sacrifical layer—ie, thin film of Ti or Au. The metal firm serves as a seed layer for electroplating and facilitates removal of the pdms membrane from the wafer 2. *note need a 2 mm gap at edge of wafer to prevent lift-off of pdms layer during processing 3. Spin thick photoresist (30um) 4. Expose so that elecrodes are raised features 5. Spin PDMS at spin rate to keep membrane thickness below pattern height 6. cure pdms Clean off any pdms adhered to electrodes 7. Electoplate metal electrodes, strain gauges, and contact pads-Pt or Au 8. O2 plasma-oxidize PDMS surface, allows PR to wet the PDMS surface, eliminating beading and ensuring a smooth, uniform coat of resist. 9. Spin resist 10. Expose area for traces. 11. Re-plasma 12. ITO adhesion layer (20nm) 13. E-beam evaporate gold film (150nm) 14. PR removal-acetone lift-off 15. Ethanol rinse and dry to prepare for passivation 16. O2 plasma 17. Spin PDMS membrane 18. Cure PDMS on top of glass substrate Channels to apply vacuum pressure to PDMS membrane Glass Glass Exposed SU-8 Unexposed SU-8

Fabrication: SU-8 Processing Step 3: Spin 2nd layer SU-8 (100μm thick), prebake, expose, postbake Equipment: Spin coater, hot plate, exposer Location: MERL Handle wafer is Au-coated Si wafer 1. Si wafer Sacrifical layer—ie, thin film of Ti or Au. The metal firm serves as a seed layer for electroplating and facilitates removal of the pdms membrane from the wafer 2. *note need a 2 mm gap at edge of wafer to prevent lift-off of pdms layer during processing 3. Spin thick photoresist (30um) 4. Expose so that elecrodes are raised features 5. Spin PDMS at spin rate to keep membrane thickness below pattern height 6. cure pdms Clean off any pdms adhered to electrodes 7. Electoplate metal electrodes, strain gauges, and contact pads-Pt or Au 8. O2 plasma-oxidize PDMS surface, allows PR to wet the PDMS surface, eliminating beading and ensuring a smooth, uniform coat of resist. 9. Spin resist 10. Expose area for traces. 11. Re-plasma 12. ITO adhesion layer (20nm) 13. E-beam evaporate gold film (150nm) 14. PR removal-acetone lift-off 15. Ethanol rinse and dry to prepare for passivation 16. O2 plasma 17. Spin PDMS membrane 18. Cure PDMS on top of glass substrate Glass Exposed SU-8 Unexposed SU-8 Loading post to support PDMS membrane

Fabrication: SU-8 Processing Step 4: Spin 3rd layer SU-8 (100μm thick), prebake, expose, postbake Equipment: Spin coater, hot plate, exposer Location: MERL Handle wafer is Au-coated Si wafer 1. Si wafer Sacrifical layer—ie, thin film of Ti or Au. The metal firm serves as a seed layer for electroplating and facilitates removal of the pdms membrane from the wafer 2. *note need a 2 mm gap at edge of wafer to prevent lift-off of pdms layer during processing 3. Spin thick photoresist (30um) 4. Expose so that elecrodes are raised features 5. Spin PDMS at spin rate to keep membrane thickness below pattern height 6. cure pdms Clean off any pdms adhered to electrodes 7. Electoplate metal electrodes, strain gauges, and contact pads-Pt or Au 8. O2 plasma-oxidize PDMS surface, allows PR to wet the PDMS surface, eliminating beading and ensuring a smooth, uniform coat of resist. 9. Spin resist 10. Expose area for traces. 11. Re-plasma 12. ITO adhesion layer (20nm) 13. E-beam evaporate gold film (150nm) 14. PR removal-acetone lift-off 15. Ethanol rinse and dry to prepare for passivation 16. O2 plasma 17. Spin PDMS membrane 18. Cure PDMS on top of glass substrate 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 Handle wafer is Au-coated Si wafer 1. Si wafer Sacrifical layer—ie, thin film of Ti or Au. The metal firm serves as a seed layer for electroplating and facilitates removal of the pdms membrane from the wafer 2. *note need a 2 mm gap at edge of wafer to prevent lift-off of pdms layer during processing 3. Spin thick photoresist (30um) 4. Expose so that elecrodes are raised features 5. Spin PDMS at spin rate to keep membrane thickness below pattern height 6. cure pdms Clean off any pdms adhered to electrodes 7. Electoplate metal electrodes, strain gauges, and contact pads-Pt or Au 8. O2 plasma-oxidize PDMS surface, allows PR to wet the PDMS surface, eliminating beading and ensuring a smooth, uniform coat of resist. 9. Spin resist 10. Expose area for traces. 11. Re-plasma 12. ITO adhesion layer (20nm) 13. E-beam evaporate gold film (150nm) 14. PR removal-acetone lift-off 15. Ethanol rinse and dry to prepare for passivation 16. O2 plasma 17. Spin PDMS membrane 18. Cure PDMS on top of glass substrate Glass Exposed SU-8 Unexposed SU-8

Fabrication: SU-8 Processing Step 6: Develop SU-8, IPA/DI rinse Equipment: Location: MERL Handle wafer is Au-coated Si wafer 1. Si wafer Sacrifical layer—ie, thin film of Ti or Au. The metal firm serves as a seed layer for electroplating and facilitates removal of the pdms membrane from the wafer 2. *note need a 2 mm gap at edge of wafer to prevent lift-off of pdms layer during processing 3. Spin thick photoresist (30um) 4. Expose so that elecrodes are raised features 5. Spin PDMS at spin rate to keep membrane thickness below pattern height 6. cure pdms Clean off any pdms adhered to electrodes 7. Electoplate metal electrodes, strain gauges, and contact pads-Pt or Au 8. O2 plasma-oxidize PDMS surface, allows PR to wet the PDMS surface, eliminating beading and ensuring a smooth, uniform coat of resist. 9. Spin resist 10. Expose area for traces. 11. Re-plasma 12. ITO adhesion layer (20nm) 13. E-beam evaporate gold film (150nm) 14. PR removal-acetone lift-off 15. Ethanol rinse and dry to prepare for passivation 16. O2 plasma 17. Spin PDMS membrane 18. Cure PDMS on top of glass substrate Glass Exposed SU-8

Fabrication: SU-8 Processing Step 7: Pipette tetrafluoropolymer (PS200 or T2494) to prevent PDMS membrane stiction Equipment: Location: MERL Handle wafer is Au-coated Si wafer 1. Si wafer Sacrifical layer—ie, thin film of Ti or Au. The metal firm serves as a seed layer for electroplating and facilitates removal of the pdms membrane from the wafer 2. *note need a 2 mm gap at edge of wafer to prevent lift-off of pdms layer during processing 3. Spin thick photoresist (30um) 4. Expose so that elecrodes are raised features 5. Spin PDMS at spin rate to keep membrane thickness below pattern height 6. cure pdms Clean off any pdms adhered to electrodes 7. Electoplate metal electrodes, strain gauges, and contact pads-Pt or Au 8. O2 plasma-oxidize PDMS surface, allows PR to wet the PDMS surface, eliminating beading and ensuring a smooth, uniform coat of resist. 9. Spin resist 10. Expose area for traces. 11. Re-plasma 12. ITO adhesion layer (20nm) 13. E-beam evaporate gold film (150nm) 14. PR removal-acetone lift-off 15. Ethanol rinse and dry to prepare for passivation 16. O2 plasma 17. Spin PDMS membrane 18. Cure PDMS on top of glass substrate 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 Handle wafer is Au-coated Si wafer 1. Si wafer Sacrifical layer—ie, thin film of Ti or Au. The metal firm serves as a seed layer for electroplating and facilitates removal of the pdms membrane from the wafer 2. *note need a 2 mm gap at edge of wafer to prevent lift-off of pdms layer during processing 3. Spin thick photoresist (30um) 4. Expose so that elecrodes are raised features 5. Spin PDMS at spin rate to keep membrane thickness below pattern height 6. cure pdms Clean off any pdms adhered to electrodes 7. Electoplate metal electrodes, strain gauges, and contact pads-Pt or Au 8. O2 plasma-oxidize PDMS surface, allows PR to wet the PDMS surface, eliminating beading and ensuring a smooth, uniform coat of resist. 9. Spin resist 10. Expose area for traces. 11. Re-plasma 12. ITO adhesion layer (20nm) 13. E-beam evaporate gold film (150nm) 14. PR removal-acetone lift-off 15. Ethanol rinse and dry to prepare for passivation 16. O2 plasma 17. Spin PDMS membrane 18. Cure PDMS on top of glass substrate 20μm clearance between loading post and PDMS membrane Glass/Quartz Exposed SU-8

Fabrication: PDMS Membrane Step 1: Clean 4” silicon wafers Equipment: wbnonmetal Location: SNF Handle wafer is Au-coated Si wafer 1. Si wafer Sacrifical layer—ie, thin film of Ti or Au. The metal firm serves as a seed layer for electroplating and facilitates removal of the pdms membrane from the wafer 2. *note need a 2 mm gap at edge of wafer to prevent lift-off of pdms layer during processing 3. Spin thick photoresist (30um) 4. Expose so that elecrodes are raised features 5. Spin PDMS at spin rate to keep membrane thickness below pattern height 6. cure pdms Clean off any pdms adhered to electrodes 7. Electoplate metal electrodes, strain gauges, and contact pads-Pt or Au 8. O2 plasma-oxidize PDMS surface, allows PR to wet the PDMS surface, eliminating beading and ensuring a smooth, uniform coat of resist. 9. Spin resist 10. Expose area for traces. 11. Re-plasma 12. ITO adhesion layer (20nm) 13. E-beam evaporate gold film (150nm) 14. PR removal-acetone lift-off 15. Ethanol rinse and dry to prepare for passivation 16. O2 plasma 17. Spin PDMS membrane 18. Cure PDMS on top of glass substrate 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 PAA (50 kDa) purchased from Polysciences (Warrington, PA). Sacrificial layer preparation: The PAA purchased as a 25% (w/v) solution in water was neutralized with a saturated solution of NaOH until reaching a pH of 7.5 with a pH indicator band test, and then diluted to the appropriate concentration. The silicon wafers were immersed in a 5% aqueous solution of HCl for 5 min, rinsed with deionized water, and dried with a stream of nitrogen gas. The surface of the polymeric substrates (e.g., PET) was rendered hydrophilic by a brief exposure to oxygen plasma (30 s, 18 W). Both of these treatments improved the wettability of the aqueous solutions of PAA and dextran on the substrates. The solutions of water-soluble polymer were filtered (0.45 mm or 5 mm pore size for solutions of polymer with less or more than 5% (w/v), respectively) and dispensed onto the substrate until about 90% of the surface was covered with the solution. The sacrificial layer was then prepared by spin-coating the substrate at 1000–4000 rpm for 15 s, and baking the film on a hot plate (at 150C for silicon, or 95C for polymeric substrates) for 2 min. Film-thickness measurements: We dissolved the sacrificial layer from over about half of the surface of the substrate with a stream of water (from a water bottle), and dried the substratewith a stream of nitrogen gas. The thickness of the film was then determined by averaging profilometry measurements at three different locations on each substrate. Microfabrication: We prepared the photoresist structures according to the manufacturer’s instructions. For the characterization of the etching speed of PAA and dextran, for the experiments with Ni electrodeposition, and for the shadow-mask evaporation of metals, we used sacrificial layers prepared from solutions of polymer of 5% (w/v) and spin-coated at 3000 rpm. The sacrificial layers for the free-standing structures were prepared from solutions of 19% PAA and 20% dextran (w/v) and spin-coated at 1000 rpm. The characterization of the etching of the sacrificial layer was carried out with deionized water. For all other experiments we added Tween 20, at a concentration of 0.05%, to improve the wettability of the water (or NaCl solution) on the SU-8 features. The nickel was electrodeposited at constant current, between 1 and 20 mAcm2. The free-standing features were released by immersion in water for 40 s. Silicon PAA

Sacrificial Layers—PDMS Micromachining 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 Water-Soluble Sacrificial Layers for Surface Micromachining Vincent Linder, Byron D. Gates, Declan Ryan, Babak A. Parviz, and George M. Whitesides* SMALL 2005

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 2mm gap at edge of wafer to prevent lift-off of PDMS during processing Handle wafer is Au-coated Si wafer 1. Si wafer Sacrifical layer—ie, thin film of Ti or Au. The metal firm serves as a seed layer for electroplating and facilitates removal of the pdms membrane from the wafer 2. *note need a 2 mm gap at edge of wafer to prevent lift-off of pdms layer during processing 3. Spin thick photoresist (30um) 4. Expose so that elecrodes are raised features 5. Spin PDMS at spin rate to keep membrane thickness below pattern height 6. cure pdms Clean off any pdms adhered to electrodes 7. Electoplate metal electrodes, strain gauges, and contact pads-Pt or Au 8. O2 plasma-oxidize PDMS surface, allows PR to wet the PDMS surface, eliminating beading and ensuring a smooth, uniform coat of resist. 9. Spin resist 10. Expose area for traces. 11. Re-plasma 12. ITO adhesion layer (20nm) 13. E-beam evaporate gold film (150nm) 14. PR removal-acetone lift-off 15. Ethanol rinse and dry to prepare for passivation 16. O2 plasma 17. Spin PDMS membrane 18. Cure PDMS on top of glass substrate Silicon PAA PDMS

Fabrication: Electrode Array Step 4: Align beryllium copper shadow mask and temporarily bond. Equipment: EV aligner Location: SNF 30μm width tracks for electrode connections Handle wafer is Au-coated Si wafer 1. Si wafer Sacrifical layer—ie, thin film of Ti or Au. The metal firm serves as a seed layer for electroplating and facilitates removal of the pdms membrane from the wafer 2. *note need a 2 mm gap at edge of wafer to prevent lift-off of pdms layer during processing 3. Spin thick photoresist (30um) 4. Expose so that elecrodes are raised features 5. Spin PDMS at spin rate to keep membrane thickness below pattern height 6. cure pdms Clean off any pdms adhered to electrodes 7. Electoplate metal electrodes, strain gauges, and contact pads-Pt or Au 8. O2 plasma-oxidize PDMS surface, allows PR to wet the PDMS surface, eliminating beading and ensuring a smooth, uniform coat of resist. 9. Spin resist 10. Expose area for traces. 11. Re-plasma 12. ITO adhesion layer (20nm) 13. E-beam evaporate gold film (150nm) 14. PR removal-acetone lift-off 15. Ethanol rinse and dry to prepare for passivation 16. O2 plasma 17. Spin PDMS membrane 18. Cure PDMS on top of glass substrate Silicon PAA PDMS Shadow Mask Ti Au 20μm diameter electrodes

Fabrication: Electrode Array Step 5: Evaporate Ti adhesion layer (10nm thick) and Au layer (100nm thick) Equipment: Innotec Location: SNF 30μm width tracks for electrode connections Handle wafer is Au-coated Si wafer 1. Si wafer Sacrifical layer—ie, thin film of Ti or Au. The metal firm serves as a seed layer for electroplating and facilitates removal of the pdms membrane from the wafer 2. *note need a 2 mm gap at edge of wafer to prevent lift-off of pdms layer during processing 3. Spin thick photoresist (30um) 4. Expose so that elecrodes are raised features 5. Spin PDMS at spin rate to keep membrane thickness below pattern height 6. cure pdms Clean off any pdms adhered to electrodes 7. Electoplate metal electrodes, strain gauges, and contact pads-Pt or Au 8. O2 plasma-oxidize PDMS surface, allows PR to wet the PDMS surface, eliminating beading and ensuring a smooth, uniform coat of resist. 9. Spin resist 10. Expose area for traces. 11. Re-plasma 12. ITO adhesion layer (20nm) 13. E-beam evaporate gold film (150nm) 14. PR removal-acetone lift-off 15. Ethanol rinse and dry to prepare for passivation 16. O2 plasma 17. Spin PDMS membrane 18. Cure PDMS on top of glass substrate Silicon PAA PDMS Shadow Mask Ti Au 20μm diameter electrodes

Fabrication: Electrode Array Step 6: Remove shadow mask, O2 plasma Equipment: Drytek Location: SNF Handle wafer is Au-coated Si wafer 1. Si wafer Sacrifical layer—ie, thin film of Ti or Au. The metal firm serves as a seed layer for electroplating and facilitates removal of the pdms membrane from the wafer 2. *note need a 2 mm gap at edge of wafer to prevent lift-off of pdms layer during processing 3. Spin thick photoresist (30um) 4. Expose so that elecrodes are raised features 5. Spin PDMS at spin rate to keep membrane thickness below pattern height 6. cure pdms Clean off any pdms adhered to electrodes 7. Electoplate metal electrodes, strain gauges, and contact pads-Pt or Au 8. O2 plasma-oxidize PDMS surface, allows PR to wet the PDMS surface, eliminating beading and ensuring a smooth, uniform coat of resist. 9. Spin resist 10. Expose area for traces. 11. Re-plasma 12. ITO adhesion layer (20nm) 13. E-beam evaporate gold film (150nm) 14. PR removal-acetone lift-off 15. Ethanol rinse and dry to prepare for passivation 16. O2 plasma 17. Spin PDMS membrane 18. Cure PDMS on top of glass substrate 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 Handle wafer is Au-coated Si wafer 1. Si wafer Sacrifical layer—ie, thin film of Ti or Au. The metal firm serves as a seed layer for electroplating and facilitates removal of the pdms membrane from the wafer 2. *note need a 2 mm gap at edge of wafer to prevent lift-off of pdms layer during processing 3. Spin thick photoresist (30um) 4. Expose so that elecrodes are raised features 5. Spin PDMS at spin rate to keep membrane thickness below pattern height 6. cure pdms Clean off any pdms adhered to electrodes 7. Electoplate metal electrodes, strain gauges, and contact pads-Pt or Au 8. O2 plasma-oxidize PDMS surface, allows PR to wet the PDMS surface, eliminating beading and ensuring a smooth, uniform coat of resist. 9. Spin resist 10. Expose area for traces. 11. Re-plasma 12. ITO adhesion layer (20nm) 13. E-beam evaporate gold film (150nm) 14. PR removal-acetone lift-off 15. Ethanol rinse and dry to prepare for passivation 16. O2 plasma 17. Spin PDMS membrane 18. Cure PDMS on top of glass substrate Silicon PAA PDMS Shadow Mask Ti Au PPS *Proximity exposure **Need to characterize in BlueM Oven

Fabrication: Electrode Array Step 8: Dissolve sacrificial layer PAA in water Equipment: wbgeneral Location: SNF Handle wafer is Au-coated Si wafer 1. Si wafer Sacrifical layer—ie, thin film of Ti or Au. The metal firm serves as a seed layer for electroplating and facilitates removal of the pdms membrane from the wafer 2. *note need a 2 mm gap at edge of wafer to prevent lift-off of pdms layer during processing 3. Spin thick photoresist (30um) 4. Expose so that elecrodes are raised features 5. Spin PDMS at spin rate to keep membrane thickness below pattern height 6. cure pdms Clean off any pdms adhered to electrodes 7. Electoplate metal electrodes, strain gauges, and contact pads-Pt or Au 8. O2 plasma-oxidize PDMS surface, allows PR to wet the PDMS surface, eliminating beading and ensuring a smooth, uniform coat of resist. 9. Spin resist 10. Expose area for traces. 11. Re-plasma 12. ITO adhesion layer (20nm) 13. E-beam evaporate gold film (150nm) 14. PR removal-acetone lift-off 15. Ethanol rinse and dry to prepare for passivation 16. O2 plasma 17. Spin PDMS membrane 18. Cure PDMS on top of glass substrate Silicon PAA PDMS Shadow Mask Ti Au PPS

Fabrication: Electrode Array Step 9: Air dry device Equipment: N2 gun Handle wafer is Au-coated Si wafer 1. Si wafer Sacrifical layer—ie, thin film of Ti or Au. The metal firm serves as a seed layer for electroplating and facilitates removal of the pdms membrane from the wafer 2. *note need a 2 mm gap at edge of wafer to prevent lift-off of pdms layer during processing 3. Spin thick photoresist (30um) 4. Expose so that elecrodes are raised features 5. Spin PDMS at spin rate to keep membrane thickness below pattern height 6. cure pdms Clean off any pdms adhered to electrodes 7. Electoplate metal electrodes, strain gauges, and contact pads-Pt or Au 8. O2 plasma-oxidize PDMS surface, allows PR to wet the PDMS surface, eliminating beading and ensuring a smooth, uniform coat of resist. 9. Spin resist 10. Expose area for traces. 11. Re-plasma 12. ITO adhesion layer (20nm) 13. E-beam evaporate gold film (150nm) 14. PR removal-acetone lift-off 15. Ethanol rinse and dry to prepare for passivation 16. O2 plasma 17. Spin PDMS membrane 18. Cure PDMS on top of glass substrate Silicon PAA PDMS Shadow Mask Ti Au PPS

Fabrication: Assembly Step 1: O2 plasma PDMS and quartz surfaces Equipment: Drytek Handle wafer is Au-coated Si wafer 1. Si wafer Sacrifical layer—ie, thin film of Ti or Au. The metal firm serves as a seed layer for electroplating and facilitates removal of the pdms membrane from the wafer 2. *note need a 2 mm gap at edge of wafer to prevent lift-off of pdms layer during processing 3. Spin thick photoresist (30um) 4. Expose so that elecrodes are raised features 5. Spin PDMS at spin rate to keep membrane thickness below pattern height 6. cure pdms Clean off any pdms adhered to electrodes 7. Electoplate metal electrodes, strain gauges, and contact pads-Pt or Au 8. O2 plasma-oxidize PDMS surface, allows PR to wet the PDMS surface, eliminating beading and ensuring a smooth, uniform coat of resist. 9. Spin resist 10. Expose area for traces. 11. Re-plasma 12. ITO adhesion layer (20nm) 13. E-beam evaporate gold film (150nm) 14. PR removal-acetone lift-off 15. Ethanol rinse and dry to prepare for passivation 16. O2 plasma 17. Spin PDMS membrane 18. Cure PDMS on top of glass substrate Silicon PDMS PPS Ti Au Glass/Quartz SU-8

Fabrication: Assembly Step 2: Bond PDMS membrane to glass Handle wafer is Au-coated Si wafer 1. Si wafer Sacrifical layer—ie, thin film of Ti or Au. The metal firm serves as a seed layer for electroplating and facilitates removal of the pdms membrane from the wafer 2. *note need a 2 mm gap at edge of wafer to prevent lift-off of pdms layer during processing 3. Spin thick photoresist (30um) 4. Expose so that elecrodes are raised features 5. Spin PDMS at spin rate to keep membrane thickness below pattern height 6. cure pdms Clean off any pdms adhered to electrodes 7. Electoplate metal electrodes, strain gauges, and contact pads-Pt or Au 8. O2 plasma-oxidize PDMS surface, allows PR to wet the PDMS surface, eliminating beading and ensuring a smooth, uniform coat of resist. 9. Spin resist 10. Expose area for traces. 11. Re-plasma 12. ITO adhesion layer (20nm) 13. E-beam evaporate gold film (150nm) 14. PR removal-acetone lift-off 15. Ethanol rinse and dry to prepare for passivation 16. O2 plasma 17. Spin PDMS membrane 18. Cure PDMS on top of glass substrate Ti Au SU-8 Glass/Quartz PDMS PPS

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

Sacrificial Layers—PDMS Micromachining 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. PAA (50 kDa) purchased from Polysciences (Warrington, PA). Sacrificial layer preparation: The PAA purchased as a 25% (w/v) solution in water was neutralized with a saturated solution of NaOH until reaching a pH of 7.5 with a pH indicator band test, and then diluted to the appropriate concentration. The silicon wafers were immersed in a 5% aqueous solution of HCl for 5 min, rinsed with deionized water, and dried with a stream of nitrogen gas. The surface of the polymeric substrates (e.g., PET) was rendered hydrophilic by a brief exposure to oxygen plasma (30 s, 18 W). Both of these treatments improved the wettability of the aqueous solutions of PAA and dextran on the substrates. The solutions of water-soluble polymer were filtered (0.45 mm or 5 mm pore size for solutions of polymer with less or more than 5% (w/v), respectively) and dispensed onto the substrate until about 90% of the surface was covered with the solution. The sacrificial layer was then prepared by spin-coating the substrate at 1000–4000 rpm for 15 s, and baking the film on a hot plate (at 150C for silicon, or 95C for polymeric substrates) for 2 min. Film-thickness measurements: We dissolved the sacrificial layer from over about half of the surface of the substrate with a stream of water (from a water bottle), and dried the substratewith a stream of nitrogen gas. The thickness of the film was then determined by averaging profilometry measurements at three different locations on each substrate. Microfabrication: We prepared the photoresist structures according to the manufacturer’s instructions. For the characterization of the etching speed of PAA and dextran, for the experiments with Ni electrodeposition, and for the shadow-mask evaporation of metals, we used sacrificial layers prepared from solutions of polymer of 5% (w/v) and spin-coated at 3000 rpm. The sacrificial layers for the free-standing structures were prepared from solutions of 19% PAA and 20% dextran (w/v) and spin-coated at 1000 rpm. The characterization of the etching of the sacrificial layer was carried out with deionized water. For all other experiments we added Tween 20, at a concentration of 0.05%, to improve the wettability of the water (or NaCl solution) on the SU-8 features. The nickel was electrodeposited at constant current, between 1 and 20 mAcm2. The free-standing features were released by immersion in water for 40 s.

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)