Kadilak Andrea L, Rehaag Jessica C, Harrington Cameron A, Shor Leslie M
Department of Chemical and Biomolecular Engineering, University of Connecticut, Storrs, Connecticut 06269-3222, USA.
Biomicrofluidics. 2017 Oct 2;11(5):054109. doi: 10.1063/1.5003477. eCollection 2017 Sep.
Additive manufacturing, or 3D-printing techniques have recently begun to enable simpler, faster, and cheaper production of millifluidic devices at resolutions approaching 100-200 m. At this resolution, cell culture devices can be constructed that more accurately replicate natural environments compared with conventional culturing techniques. A number of microfluidics researchers have begun incorporating additive manufacturing into their work, using 3D-printed devices in a wide array of chemical, fluidic, and even some biological applications. Here, we describe a 3D-printed cell culture platform and demonstrate its use in culturing KT2440 bacteria for 44 h under a differential substrate gradient. Polyethylene glycol diacrylate (PEGDA) hydrogel barriers are patterned within a 3D-printed channel. Transport of the toluidine blue tracer dye through the hydrogel barriers is characterized. Nutrients and oxygen were delivered to cells in the culture region by diffusion through the PEGDA hydrogel barriers from adjacent media or saline perfusion channels. Expression of green fluorescent protein by KT2440 enabled real time visualization of cell density within the 3D-printed channel, and demonstrated cells were actively expressing protein over the course of the experiment. Cells were observed clustering near hydrogel barrier boundaries where fresh substrate and oxygen were being delivered diffusive transport, but cells were unable to penetrate the barrier. The device described here provides a versatile and easy to implement platform for cell culture in readily controlled gradient microenvironments. By adjusting device geometry and hydrogel properties, this platform could be further customized for a wide variety of biological applications.
增材制造或3D打印技术最近已开始能够以接近100-200微米的分辨率更简单、快速且廉价地生产微流控设备。在此分辨率下，可以构建细胞培养设备，与传统培养技术相比，该设备能更准确地复制自然环境。许多微流控研究人员已开始将增材制造纳入其工作中，在广泛的化学、流体甚至一些生物应用中使用3D打印设备。在此，我们描述了一种3D打印的细胞培养平台，并展示了其在差分底物梯度下培养KT2440细菌44小时的用途。聚乙二醇二丙烯酸酯（PEGDA）水凝胶屏障在3D打印通道内形成图案。对甲苯胺蓝示踪染料通过水凝胶屏障的传输进行了表征。营养物质和氧气通过从相邻培养基或盐水灌注通道扩散穿过PEGDA水凝胶屏障传递到培养区域的细胞中。KT2440表达绿色荧光蛋白能够实时可视化3D打印通道内的细胞密度，并证明细胞在实验过程中积极表达蛋白质。观察到细胞聚集在水凝胶屏障边界附近，新鲜底物和氧气通过扩散传输在此处传递，但细胞无法穿透屏障。这里描述的设备为在易于控制的梯度微环境中进行细胞培养提供了一个通用且易于实施的平台。通过调整设备几何形状和水凝胶特性，该平台可进一步针对各种生物应用进行定制。
