# Additive Manufacturing of Bone Scaffolds Using PolyJet and Stereolithography Techniques
## 2 Materials and Methods
### Permeability Analysis of 3D-Printed Scaffold Structures
#### 2.2.3 Experimental Measurements of Permeability  
To measure intrinsic permeability, a constant head permeability test setup was designed and developed, as shown in Figure 2. This setup consisted of a fluid holding tank with a 20 L capacity maintained at 800 mm above the surface of the scaffold to achieve hydrostatic pressure on the scaffold surface. A clamping device was required that could hold the scaffold structures for the flow rate measurements. For this purpose, a stainless steel scaffold holding clamp and O-rings were designed according to the dimensions of the scaffold structures using Creo Parametric 4.0, which were machined using a high-precision CNC machine. To validate the clamp performance and tightness, a leak-tight test was performed on the clamp. For this test, a blind sample with similar dimensions to the scaffold dimensions was designed and three copies of the model were printed using the Polylet printer. The leak-tight test was performed with tap water on the three blind samples under same conditions at which the permeability tests were carried out. The leak-tight test was performed for three hours and no leakages in the clamp were observed. The leak-tight test was performed after every few runs to check that the O-rings were still good and not flattened or damaged after several work cycles. The temperature of the laboratory was maintained at 22 \({}^{\circ}\)C as a requirement for other instruments in the lab. The ambience can affect the sample, test fluids, and sealants. Three different test fluids, including the tap water, 15% glycerol-water solution, and 20% glycerol-water solutions were selected to measure the permeability of the 3D-printed scaffold samples. These test fluids have also been used by others researchers for permeability analyses of different scaffold structures [23]. In previous studies, raw seed oil [39], olive oil [40], acrylic cement [41], silicone oil [14], and corn oil [42] were also used as test fluids. The purpose of using three different test fluid was to observe the effects of the viscosity of each fluid on the permeability. The two different water-glycerol solutions were prepared to simulate the viscosity of human blood without cells ranging from \(1\times 10^{-3}\) Pa.s to \(1.3\times 10^{-3}\) Pa.s [43]. For test fluids, glycerol was purchased from Alfa Aesar England. The viscosity of the prepared test fluids was measured with the help of the Physica MCR-301 Rheometer-Anton Paar instrument. Three samples of 3 mL were tested to measure the viscosity values for each test fluid. Every reading was taken for the 100 th iteration and the obtained average viscosity levels of all fluids were about \(1\times 10^{-3}\) Pa.s, \(2.35\times 10^{-3}\) Pa.s, and \(2.87\times 10^{-3}\) Pa.s for tap water, 15% glycerol-water, and 20% glycerol-water solutions, respectively, at 22 \({}^{\circ}\)C. The flow rate was measured by recording the time required for 500 mL to pass through the scaffold structures in only one direction (i.e., the sample-built direction). The cubic scaffold structures were omnidirectional and were, therefore, tested only in one direction. The hexagonal closed packed structures contained only one primary axis with less obstruction and were tested along this direction (built direction); thus, three readings of the times required for 500 mL of fluid to pass through each scaffold sample were recorded to obtain the average value, in order to ensure better flow rate results. The same procedure was repeated for all the test fluids.