## 3 Results  
In this study, we developed a method to print hydrogel scaffolds using various combinations of 3 BE and dispensing the stainless steel needle head through a self-fabricated 3D bioprinting system. First of all, we printed the combinations of 3 BE-1 with different needle heads as a straight-line. It was found that the 3 BE-1 with larger diameter of needle heads exhibited an irregular edge in the printed line as indicated by the arrows (Figure 1). However, the 3 BE-2 or 3 BE-3 with larger diameter of needle head presented a uniform edge in the printed line. Then, we tested the quality of line width after the printing process under fixed write height, extrudate, and lift nozzle. Analytical results revealed that both needle sizes and 3 BE concentrations affect the quality of the printed line. Among all combinationsof printing parameters, it was found that the combination of 3 BE-2 with all diameter of needle sizes produced smooth and regular line widths (Figure 2). Similar results can also be discovered in the combinations of 3 BE-3. Hence, these findings reflected that both 3 BE-2 and 3 BE-3 combinations possessed great potential to print a stable construct. Based on the abovementioned printing parameters, we then printed a more complex 3D cubic-like construct using the combinations of 3 BE-2 with different needle heads. Figure 3a depicts the 3D cubic-like construct successfully printed under the combination of 3 BE-2 with \(\Phi\) 0.13 mm needle head. Nevertheless, we found that the generated 3D cubic-like structure collapsed after 4 h of printing even those printed samples with larger needle heads. Thus, we attempted to test the combinations of 3 BE-3 according to the parameters as shown in Table 1. From this experiment, we discovered that a stable and smooth 3D cubic-like construct can be printed using the combinations of 3 BE-3 (Figure 3b). Interestingly, the shape of the printed construct can be maintained for over 24 h post-printing. Figure 1: The combinations of 3 BE-1 with different diameters of stainless steel needle heads were used to print a straight-line: (a) \(\Phi\) 0.13 mm, (b) \(\Phi\) 0.33 mm, and (c) \(\Phi\) 0.9 mm. Figure 3: The 3D cubic-like constructs were printed through the self-fabricated 3D bioprinting system: (a) 3 BE-2 combined with \(\Phi\) 0.13 mm needle head and (b) 3 BE-3 combined with \(\Phi\) 0.13 mm needle head. The shape of the printed structure can be maintained for over 24 h using the combinations of 3 BE-3. Figure 2: The printed results from the self-fabricated 3D bioprinting system using the combinations of 3 BE-2 with (a) \(\Phi\) 0.13 mm, (b) \(\Phi\) 0.33 mm, and (c) \(\Phi\) 0.9 mm needle head. In order to evaluate the biocompatibility of 3 BE for cell encapsulation in 3D printing applications, firstly, we tested the cell viability of L929 fibroblast within various concentrations of 3 BE in 2D monolayer. The live/dead analysis showed that more live cells were found in all 3 BE groups than the dead cells (Figure 4). Among all formulations of 3 BE hydrogel, the highest amount of live cells were seen in 3 BE-1. In addition, we found that the cell number was rather decreased as the concentration of the 3 BE increased. To confirm this result, we performed the quantitative analysis of cell viability cultured with various concentrations of 3 BE shown in Figure 5. Consistent with live/dead assay, MTT analysis showed that 3 BE-1 presented the highest cell viability compared to other 3 BE groups and control. Even though the cell viability was decreased in 3 BE-2 and 3 BE-3, the number of viable cells was still above 70%, indicating the promising effect of 3 BE for cell encapsulation. Based on the printability and biocompatibility results evaluation, it suggested that the ideal concentration as a bioink was 3 BE-3. This concentration is further adopted for the bioprinting experiments. After establishing idea concentration of 3 BE hydrogel, we then used this concentration to print cell-laden hydrogels.