3. NucBlue™
Fixed
Cell
ReadyProbes™
Reagent
(ThermoFisher).
4. AlexaFluor
®
488
Phalloidin
(AF488-Phalloidin;
ThermoFisher).
5. CellTracker™ Red CMTPX Dye (ThermoFisher).
6. Fixative solution: 3.2% Paraformaldehyde solution prepared by
diluting 32% Paraformaldehyde aqueous solution 1:9 with
Hank’s Buffered Saline Solution supplemented with calcium
and magnesium (HBSS +/+).
7. Staining solution: Prior to imaging, prepare a fresh stock solution with NucBlue (2 drops per mL) and AF488-Phalloidin
(5 μg/mL) in HBSS +/+.
3 Methods
3.1 Scaffold Design
Different options are currently available for generating a suitable
3D model for AM in biomedical engineering. On one hand, the
user can design a construct using CAD software and principle
features of the target tissue. The advantages of this method are
the freedom of designing geometries with specific features and the
ability to break down, or combine, features into multiple components. On the other hand, the user can combine high-resolution
medical images, such as computed tomography (CT), micro-CT, or
magnetic resonance imaging (MRI), with CAD to design a construct informed by anatomical information about the native tissue
at the implant site. The generation of a 3D model from medical
images requires post-processing and usually starts with image segmentation [8]. In this step, the volumetric image data can be
filtered and partitioned into multiple segments, or pixels, which
define the boundaries and spatial arrangement of objects in the
image. Then, the volume representation of the 3D image can be
applied for volume rendering and generation of anatomic models
[9]. Moreover, the segmented objects can be used to create a 3D
triangle mesh, in which triangular facets approximate the external
shape of the object (see Note 10). One of the main advantages of
having a 3D image containing labeled objects is the ease of engineering a 3D model with specific anisotropy and mechanical properties that can be useful to predict its mechanical deformation using
finite element (FE) simulation.
In this description of the method, we employ CAD software to
design tubular constructs without medical imaging:
1. Draw the tubular scaffolds in AutoCAD
® software (Autodesk,
San Rafael, CA, USA) with internal diameter 10 mm and
external diameter 12 mm and longitudinal length 30 mm
with two different window geometries (Fig. 1):
Surface Tension-Assisted Additive Manufacturing
153
Fixed
Cell
ReadyProbes™
Reagent
(ThermoFisher).
4. AlexaFluor
®
488
Phalloidin
(AF488-Phalloidin;
ThermoFisher).
5. CellTracker™ Red CMTPX Dye (ThermoFisher).
6. Fixative solution: 3.2% Paraformaldehyde solution prepared by
diluting 32% Paraformaldehyde aqueous solution 1:9 with
Hank’s Buffered Saline Solution supplemented with calcium
and magnesium (HBSS +/+).
7. Staining solution: Prior to imaging, prepare a fresh stock solution with NucBlue (2 drops per mL) and AF488-Phalloidin
(5 μg/mL) in HBSS +/+.
3 Methods
3.1 Scaffold Design
Different options are currently available for generating a suitable
3D model for AM in biomedical engineering. On one hand, the
user can design a construct using CAD software and principle
features of the target tissue. The advantages of this method are
the freedom of designing geometries with specific features and the
ability to break down, or combine, features into multiple components. On the other hand, the user can combine high-resolution
medical images, such as computed tomography (CT), micro-CT, or
magnetic resonance imaging (MRI), with CAD to design a construct informed by anatomical information about the native tissue
at the implant site. The generation of a 3D model from medical
images requires post-processing and usually starts with image segmentation [8]. In this step, the volumetric image data can be
filtered and partitioned into multiple segments, or pixels, which
define the boundaries and spatial arrangement of objects in the
image. Then, the volume representation of the 3D image can be
applied for volume rendering and generation of anatomic models
[9]. Moreover, the segmented objects can be used to create a 3D
triangle mesh, in which triangular facets approximate the external
shape of the object (see Note 10). One of the main advantages of
having a 3D image containing labeled objects is the ease of engineering a 3D model with specific anisotropy and mechanical properties that can be useful to predict its mechanical deformation using
finite element (FE) simulation.
In this description of the method, we employ CAD software to
design tubular constructs without medical imaging:
1. Draw the tubular scaffolds in AutoCAD
® software (Autodesk,
San Rafael, CA, USA) with internal diameter 10 mm and
external diameter 12 mm and longitudinal length 30 mm
with two different window geometries (Fig. 1):
Surface Tension-Assisted Additive Manufacturing
153