首页> 美国卫生研究院文献>Journal of Visualized Experiments : JoVE >Printing Thermoresponsive Reverse Molds for the Creation of Patterned Two-component Hydrogels for 3D Cell Culture
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Printing Thermoresponsive Reverse Molds for the Creation of Patterned Two-component Hydrogels for 3D Cell Culture

机译:印刷用于制造3D细胞培养的图案化双组分水凝胶的热敏反型模具

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摘要

Bioprinting is an emerging technology that has its origins in the rapid prototyping industry. The different printing processes can be divided into contact bioprinting1-4 (extrusion, dip pen and soft lithography), contactless bioprinting5-7 (laser forward transfer, ink-jet deposition) and laser based techniques such as two photon photopolymerization8. It can be used for many applications such as tissue engineering9-13, biosensor microfabrication14-16 and as a tool to answer basic biological questions such as influences of co-culturing of different cell types17. 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 solutions18. 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.
机译:生物打印是一种新兴技术,其起源于快速原型制作行业。不同的印刷工艺可以分为接触式生物印刷 1-4 (挤压,蘸水笔和软光刻),非接触式生物印刷 5-7 (激光正向转印,喷墨)沉积)和基于激光的技术,例如两次光子光聚合 8 。它可用于许多应用,例如组织工程 9-13 ,生物传感器微加工 14-16 ,并可作为回答基本生物学问题(例如共培养影响)的工具不同的电池类型 17 。与普通的光刻或软光刻方法不同,挤出生物打印的优点是不需要单独的掩模或印模。使用CAD软件,可以根据操作员的要求快速更改和调整结构设计。与基于光刻的方法相比,这使生物打印更具灵活性。在此,我们演示了牺牲模具的打印,以使用水凝胶内的多个支柱为例,创建多材料3D结构。这些支柱可以代表用于神经网络导管内的血管网络或管的空心结构。用于牺牲性模具的材料是poloxamer 407,它是一种具有出色印刷性能的热响应性聚合物,在4°C时为液体,在高于其胶凝温度〜20°C的情况下,对于24.5%w / v溶液为固体 18 。该性质使得可以根据需要洗脱基于泊洛沙姆的牺牲模,并且相对于固体材料的缓慢溶解(特别是对于狭窄的几何形状)而言具有优势。将泊洛沙姆印在显微镜载玻片上以创建牺牲模具。将琼脂糖移入模具中并冷却直至胶凝。在冰冷的水中洗脱泊洛沙姆后,琼脂糖模具中的空隙中充满了掺有FITC标记的纤维蛋白原的海藻酸甲酯。然后将填充的空隙与UV交联,并用落射荧光显微镜对构建体成像。

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