Department of Bioengineering, University of Pennsylvania, Philadelphia, PA, USA.
Department of Cell & Developmental Biology, University of Pennsylvania, Philadelphia, PA, USA.
Methods Mol Biol. 2024;2805:31-50. doi: 10.1007/978-1-0716-3854-5_3.
Cell patterning for 3D culture has increased our understanding of how cells interact among themselves and with their environment during tissue morphogenesis. Building cell communities from the bottom up with size and compositional control is invaluable for studies of morphological transitions. Here, we detail Photolithographic DNA-programmed Assembly of Cells (pDPAC). pDPAC uses a photoactive polyacrylamide gel substrate to capture single-stranded DNA on a 2D surface in large-scale, highly resolved patterns using the photomask technology. Cells are then functionalized with a complementary DNA strand, enabling cells to be temporarily adhered to distinct locations only where their complementary strand is patterned. These temporary 2D patterns can be transferred to extracellular matrix hydrogels for 3D culture of cells in biomimetic microenvironments. Use of a polyacrylamide substrate has advantages, including a simpler photolithography workflow, lower non-specific cell adhesion, and lower stiction to ECM hydrogels during release of patterned hydrogels. The protocol is equally applicable to large (cm)-scale patterns and repetitive arrays of smaller-scale cell interaction or migration experiments.
细胞图案化的 3D 培养增加了我们对细胞在组织形态发生过程中如何相互作用以及与环境相互作用的理解。通过自下而上的方法构建具有大小和组成控制的细胞群落，对于形态转变的研究是非常有价值的。在这里，我们详细介绍了光聚合 DNA 编程细胞组装（pDPAC）。pDPAC 使用光活性聚丙烯酰胺凝胶基底，使用光掩模技术在 2D 表面上以大规模、高分辨率的图案捕获单链 DNA。然后，用互补的 DNA 链对细胞进行功能化，使细胞只能在与其互补链图案化的特定位置暂时黏附。这些临时的 2D 图案可以转移到细胞外基质水凝胶中，用于在仿生微环境中培养细胞。使用聚丙烯酰胺基底具有优势，包括更简单的光刻工作流程、更低的非特异性细胞黏附以及在释放图案化水凝胶时与 ECM 水凝胶的更低的黏附力。该方案同样适用于大（cm）规模的图案以及较小规模的细胞相互作用或迁移实验的重复阵列。
