substrate and each initiator (see Note 3 for the selection of
initiators). Use Scotch tape to fix the initiators.
7. Place the collecting platform onto the stage. Check the collecting substrates and initiators are fixed during the stage motion
(see Note 4). Use scotch tape to secure if necessary.
8. Adjust the position of the initiators and the collecting substrate. Tune the stage motion using LabVIEW. Ensure the
two initiators move beneath the metallic tip alternatively during stage movement.
9. Manually adjust the height of the z-stage. The distance
between the metallic tip and the initiator should be
0.05–0.5 mm. Figure 2 shows the LEP configuration at this
point.
10. Start the pump. Wait till a droplet is visible at the end of the
metallic tip. Then set the stage into motion. The program
controls the stage movement, which is translated into fiber
patterns, as shown in Fig. 3a–c.
11. Switch on the voltage (see Note 5). Applied voltage ranges
from 50 V to 700 V for PS-DMF solution, from 50 V to
150 V for PEO solution, and from 100 V to 400 V for gelatin
solution (see Note 6 for controlling the fiber morphology).
12. Switch on the safety switch.
13. Wait for the desired pattern to complete. Switch off the voltage
and the safety switch.
14. Stop the stage motion using LabVIEW. Lower the stage.
Transfer the collecting substrate from the stage to a designated
petri-dish (see Note 7). The fibers should be patterned on the
substrate, as shown in Fig. 3d, e.
3.2 Cell Culture
1. Pattern gelatin fibers on a 20 Â 20 Â 0.1 mm PDMS film using
the LEP method as previously described.
2. To crosslink the gelatin fibers, immerse the specimen into a
crosslinking solution formed by 25 mM of EDC and 10 mM of
NHS in ethanol-water (9:1 v/v) [11]. Store the specimen at
4
C for 24 h to complete crosslinking.
3. Rinse the specimen with water to remove the crosslinking
reagents (see Note 8).
4. Dry the specimen in a desiccator for 24 h.
5. To sterilize the specimen for cell culture use, immerse it in 70%
ethanol for 1 h.
6. Rinse the specimen with PBS for three times. Immerse the
specimen in cell culture medium and store at 37
C.
7. Take a flask of confluent EA.hy926 cells. To resuspend the cells,
remove the medium and wash the cells with 2 mL PBS.
Low Voltage Electrospinning Patterning
131
initiators). Use Scotch tape to fix the initiators.
7. Place the collecting platform onto the stage. Check the collecting substrates and initiators are fixed during the stage motion
(see Note 4). Use scotch tape to secure if necessary.
8. Adjust the position of the initiators and the collecting substrate. Tune the stage motion using LabVIEW. Ensure the
two initiators move beneath the metallic tip alternatively during stage movement.
9. Manually adjust the height of the z-stage. The distance
between the metallic tip and the initiator should be
0.05–0.5 mm. Figure 2 shows the LEP configuration at this
point.
10. Start the pump. Wait till a droplet is visible at the end of the
metallic tip. Then set the stage into motion. The program
controls the stage movement, which is translated into fiber
patterns, as shown in Fig. 3a–c.
11. Switch on the voltage (see Note 5). Applied voltage ranges
from 50 V to 700 V for PS-DMF solution, from 50 V to
150 V for PEO solution, and from 100 V to 400 V for gelatin
solution (see Note 6 for controlling the fiber morphology).
12. Switch on the safety switch.
13. Wait for the desired pattern to complete. Switch off the voltage
and the safety switch.
14. Stop the stage motion using LabVIEW. Lower the stage.
Transfer the collecting substrate from the stage to a designated
petri-dish (see Note 7). The fibers should be patterned on the
substrate, as shown in Fig. 3d, e.
3.2 Cell Culture
1. Pattern gelatin fibers on a 20 Â 20 Â 0.1 mm PDMS film using
the LEP method as previously described.
2. To crosslink the gelatin fibers, immerse the specimen into a
crosslinking solution formed by 25 mM of EDC and 10 mM of
NHS in ethanol-water (9:1 v/v) [11]. Store the specimen at
4
C for 24 h to complete crosslinking.
3. Rinse the specimen with water to remove the crosslinking
reagents (see Note 8).
4. Dry the specimen in a desiccator for 24 h.
5. To sterilize the specimen for cell culture use, immerse it in 70%
ethanol for 1 h.
6. Rinse the specimen with PBS for three times. Immerse the
specimen in cell culture medium and store at 37
C.
7. Take a flask of confluent EA.hy926 cells. To resuspend the cells,
remove the medium and wash the cells with 2 mL PBS.
Low Voltage Electrospinning Patterning
131