3.4 SEM Analysis
To analyze the scaffold under scanning electron microscope, the
steps below should be followed (described here for a Philips XL30
SEM):
1. Soak the scaffold in liquid nitrogen for 5 min.
2. Cut the scaffold with a razor blade along the vertical axis.
3. Apply a carbon conductive tape to the sample holders, and stick
the samples on them with the section of interest facing upward.
4. Load the scaffolds on the rotating platform inside the gold
coating chamber of the sputter coating apparatus in use.
Check Note 18 for the importance of a proper coating.
5. Turn on the machine and apply vacuum.
6. Flush four times with argon.
7. Sputter coat with gold for 100 s.
The samples are then ready to be loaded in the SEM and
imaged:
1. Vent the chamber and open it.
2. Load the sample holders onto the platform and close the
chamber.
3. Apply vacuum. Follow the recommendation in Note 19 for
proper imaging.
4. Switch on the electron beam and set it at 10 kV.
5. Focus at a low magnification and then go to the desired magnification and focus again. Adjust stigmatism if required.
6. Set the working distance at the characteristic value for the
microscope in use and adjust the focus again. Repeat this step
until the actual working distance value is as close as possible to
the characteristic one.
7. Image the cross section of the scaffolds to have a clear view of
the filament diameter and of the lateral porosity. To avoid
damages to the scaffold, see Note 20.
8. Measure the filaments diameter, the filament distance, and the
layer thickness in the SEM image to characterize the scaffold.
4 Notes
1. To avoid unreliable data during DSC measurements and that
the pan sticks in the machine, ensure that the scale, the tweezers, the pans, and the lids are clean.
2. The sample should be cut so that to avoid irregularities in
shape. If the contact area between the sample and the pan is
sufficiently uniform, the heat transmission will be regular.
AM of Thermoplastics for Tissue Engineering
91
To analyze the scaffold under scanning electron microscope, the
steps below should be followed (described here for a Philips XL30
SEM):
1. Soak the scaffold in liquid nitrogen for 5 min.
2. Cut the scaffold with a razor blade along the vertical axis.
3. Apply a carbon conductive tape to the sample holders, and stick
the samples on them with the section of interest facing upward.
4. Load the scaffolds on the rotating platform inside the gold
coating chamber of the sputter coating apparatus in use.
Check Note 18 for the importance of a proper coating.
5. Turn on the machine and apply vacuum.
6. Flush four times with argon.
7. Sputter coat with gold for 100 s.
The samples are then ready to be loaded in the SEM and
imaged:
1. Vent the chamber and open it.
2. Load the sample holders onto the platform and close the
chamber.
3. Apply vacuum. Follow the recommendation in Note 19 for
proper imaging.
4. Switch on the electron beam and set it at 10 kV.
5. Focus at a low magnification and then go to the desired magnification and focus again. Adjust stigmatism if required.
6. Set the working distance at the characteristic value for the
microscope in use and adjust the focus again. Repeat this step
until the actual working distance value is as close as possible to
the characteristic one.
7. Image the cross section of the scaffolds to have a clear view of
the filament diameter and of the lateral porosity. To avoid
damages to the scaffold, see Note 20.
8. Measure the filaments diameter, the filament distance, and the
layer thickness in the SEM image to characterize the scaffold.
4 Notes
1. To avoid unreliable data during DSC measurements and that
the pan sticks in the machine, ensure that the scale, the tweezers, the pans, and the lids are clean.
2. The sample should be cut so that to avoid irregularities in
shape. If the contact area between the sample and the pan is
sufficiently uniform, the heat transmission will be regular.
AM of Thermoplastics for Tissue Engineering
91