4. Increase the temperature of the hotplate to 95
C and hold for
7 min.
5. Place the wafer and photomask in the mask aligner with the
following settings: vacuum contact mode, power 9 mW/cm
2 ,
exposure time 23 s.
6. Place the wafer on hot plate immediately after exposure at
65
C for 3 min followed by an increase to 95
C for 9 min.
7. Immerse the UV-exposed wafer in SU-8 developer for 10 min,
while gently agitating the wafer with forceps.
8. Rinse with IPA. If white residue comes appears on the wafer,
place back in the developer for additional time until no white
residue appears.
9. Dry with nitrogen gas.
10. Hard-bake the mold on a hotplate set at 120
C for 15 min.
11. Utilize the Parylene coater to deposit Parylene-C on the mold
to facilitate the release of PDMS from the mold.
3.2 Fabrication
of PDMS Stamps
(Fig. 1b)
1. Prepare a 10:1 (w/w) elastomer base to curing agent solution
of PDMS on a weighing dish.
2. For a 4
00 wafer, mix 40 g of PDMS to obtain a thickness of
~5 mm.
3. Place the PDMS in a desiccator for 30 min to remove air
bubbles from the mixture.
4. Place the mold on a flat aluminum foil sheet and wrap the foil
around the edges of the mold to make an aluminum boat to
contain the PDMS.
5. Pour the degassed PDMS on the mold and place back in the
desiccator for 15 min to remove any remaining bubbles.
6. Place the aluminum boat containing the mold and uncured
PDMS in a convection oven at 80
C for 2 h to fully cure the
PDMS. Ensure that the surface of the convection oven is level
to achieve a uniform thickness of the PDMS stamps.
7. Once the PDMS is fully cured, cut the PDMS with a razorblade
around the perimeter of the mold and remove the aluminum
foil from the mold.
8. Slowly remove the PDMS from the mold and cut it into small
1 cm  2 cm blocks; inspect pillars in microscope prior to
stamping. Note that the Parylene coating on the mold facilitates this step and helps protect the mold for multiple uses.
3.3 Stamping
Protocol (Fig. 2a)
1. Perform all the following procedures under a biological hood
using sterile techniques.
2. Place the PDMS chip in a 60-mm dish filled with 70% ethanol
(the whole chip should be covered) for 5 min.
Nanofountain Probe Electroporation for Monoclonal Cell Line Generation
63
C and hold for
7 min.
5. Place the wafer and photomask in the mask aligner with the
following settings: vacuum contact mode, power 9 mW/cm
2 ,
exposure time 23 s.
6. Place the wafer on hot plate immediately after exposure at
65
C for 3 min followed by an increase to 95
C for 9 min.
7. Immerse the UV-exposed wafer in SU-8 developer for 10 min,
while gently agitating the wafer with forceps.
8. Rinse with IPA. If white residue comes appears on the wafer,
place back in the developer for additional time until no white
residue appears.
9. Dry with nitrogen gas.
10. Hard-bake the mold on a hotplate set at 120
C for 15 min.
11. Utilize the Parylene coater to deposit Parylene-C on the mold
to facilitate the release of PDMS from the mold.
3.2 Fabrication
of PDMS Stamps
(Fig. 1b)
1. Prepare a 10:1 (w/w) elastomer base to curing agent solution
of PDMS on a weighing dish.
2. For a 4
00 wafer, mix 40 g of PDMS to obtain a thickness of
~5 mm.
3. Place the PDMS in a desiccator for 30 min to remove air
bubbles from the mixture.
4. Place the mold on a flat aluminum foil sheet and wrap the foil
around the edges of the mold to make an aluminum boat to
contain the PDMS.
5. Pour the degassed PDMS on the mold and place back in the
desiccator for 15 min to remove any remaining bubbles.
6. Place the aluminum boat containing the mold and uncured
PDMS in a convection oven at 80
C for 2 h to fully cure the
PDMS. Ensure that the surface of the convection oven is level
to achieve a uniform thickness of the PDMS stamps.
7. Once the PDMS is fully cured, cut the PDMS with a razorblade
around the perimeter of the mold and remove the aluminum
foil from the mold.
8. Slowly remove the PDMS from the mold and cut it into small
1 cm  2 cm blocks; inspect pillars in microscope prior to
stamping. Note that the Parylene coating on the mold facilitates this step and helps protect the mold for multiple uses.
3.3 Stamping
Protocol (Fig. 2a)
1. Perform all the following procedures under a biological hood
using sterile techniques.
2. Place the PDMS chip in a 60-mm dish filled with 70% ethanol
(the whole chip should be covered) for 5 min.
Nanofountain Probe Electroporation for Monoclonal Cell Line Generation
63