Müller Michael, Becher Jana, Schnabelrauch Matthias, Zenobi-Wong Marcy
Department of Health Science & Technology, Cartilage Engineering & Regeneration.
J Vis Exp. 2013 Jul 10(77):e50632. doi: 10.3791/50632.
Bioprinting is an emerging technology that has its origins in the rapid prototyping industry. The different printing processes can be divided into contact bioprinting(1-4) (extrusion, dip pen and soft lithography), contactless bioprinting(5-7) (laser forward transfer, ink-jet deposition) and laser based techniques such as two photon photopolymerization(8). It can be used for many applications such as tissue engineering(9-13), biosensor microfabrication(14-16) and as a tool to answer basic biological questions such as influences of co-culturing of different cell types(17). Unlike common photolithographic or soft-lithographic methods, extrusion bioprinting has the advantage that it does not require a separate mask or stamp. Using CAD software, the design of the structure can quickly be changed and adjusted according to the requirements of the operator. This makes bioprinting more flexible than lithography-based approaches. Here we demonstrate the printing of a sacrificial mold to create a multi-material 3D structure using an array of pillars within a hydrogel as an example. These pillars could represent hollow structures for a vascular network or the tubes within a nerve guide conduit. The material chosen for the sacrificial mold was poloxamer 407, a thermoresponsive polymer with excellent printing properties which is liquid at 4 °C and a solid above its gelation temperature ~20 °C for 24.5% w/v solutions(18). This property allows the poloxamer-based sacrificial mold to be eluted on demand and has advantages over the slow dissolution of a solid material especially for narrow geometries. Poloxamer was printed on microscope glass slides to create the sacrificial mold. Agarose was pipetted into the mold and cooled until gelation. After elution of the poloxamer in ice cold water, the voids in the agarose mold were filled with alginate methacrylate spiked with FITC labeled fibrinogen. The filled voids were then cross-linked with UV and the construct was imaged with an epi-fluorescence microscope.
生物打印是一项起源于快速成型行业的新兴技术。不同的打印工艺可分为接触式生物打印（1-4）（挤出、蘸笔和软光刻）、非接触式生物打印（5-7）（激光正向转移、喷墨沉积）以及基于激光的技术，如双光子光聚合（8）。它可用于许多应用，如组织工程（9-13）、生物传感器微制造（14-16），并作为回答基本生物学问题的工具，如不同细胞类型共培养的影响（17）。与普通光刻或软光刻方法不同，挤出式生物打印的优点是不需要单独的掩膜或印章。使用计算机辅助设计（CAD）软件，可以根据操作员的要求快速更改和调整结构设计。这使得生物打印比基于光刻的方法更加灵活。在此，我们以水凝胶内的一系列支柱为例，展示了用于创建多材料三维结构的牺牲模具的打印过程。这些支柱可以代表血管网络的中空结构或神经导管内的管道。用于牺牲模具的材料是泊洛沙姆407，这是一种具有优异打印性能的热响应聚合物，在4℃时为液体，对于24.5% w/v的溶液，在其凝胶化温度约20℃以上为固体（18）。这种特性使得基于泊洛沙姆的牺牲模具能够按需洗脱，并且相对于固体材料的缓慢溶解具有优势，特别是对于狭窄的几何形状。将泊洛沙姆打印在显微镜载玻片上以创建牺牲模具。将琼脂糖吸移到模具中并冷却直至凝胶化。在冰冷的水中洗脱泊洛沙姆后，用异硫氰酸荧光素（FITC）标记的纤维蛋白原加标的甲基丙烯酸藻酸盐填充琼脂糖模具中的空隙。然后用紫外线对填充的空隙进行交联，并用落射荧光显微镜对构建体进行成像。
