A Dual Gradient Assay for the Parametric Analysis of Cell-Surface Interactions
A Dual Gradient Assay for the Parametric Analysis of Cell-Surface Interactions
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DOI：
10.1002/smll.201200235
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发表时间：
2012-08-20
期刊：
影响因子：
13.3
通讯作者：
Gadegaard, Nikolaj
中科院分区：
文献类型：
--
作者：
Reynolds, Paul M.;Pedersen, Rasmus H.;Gadegaard, Nikolaj
细胞对微沟槽的反应是使用一种新的分析格式，包括连续变化的沟槽间距和深度的正交梯度。双层蚀刻掩模是使用微图案和等离子体聚合物沉积的组合创建的。采用光刻技术制备了沟槽宽度为8 μ m的硅衬底，沟槽宽度从8 μ m以0.5 μ m的步长增加到10 mm。等离子体聚合的己烷薄膜在这些凹槽的一端厚为120nm，另一端厚为10nm，沉积在扩散掩膜下。图案化样品的反应蚀刻将凹槽间距和凹槽深度的梯度转移到硅衬底中。使用凹槽深度梯度超过两个数量级（小于10纳米到超过1000纳米）的硅母片来创建注塑镶嵌体，以大规模复制筛选地形。聚碳酸酯复制品被模塑用于细胞培养研究，并研究了地形作为高通量筛选平台的功能。我们研究了MDCK、h-TERT成纤维细胞和LE2内皮细胞对地形线索变化的附着和形态学反应，目的是确定沟槽间距和深度的最佳组合，以引起每种细胞类型的量身定制的反应。当考虑到在单一基质上筛选的地形特征范围时，这种新的分析方法代表了生物材料界面地形线索参数化设计和分析的重要一步。
Cellular response to microgrooves is addressed using a new assay format, comprising orthogonal gradients of continuously varied groove pitch and depth. Dual layer etch masks are created using a combination of micropatterning and plasma polymer deposition. A silicon substrate with a constant groove width of 8 mu m and with ridge width increasing from 8 mu m in 0.5 mu m steps across 10 mm is fabricated by photolithography. A plasma-polymerized hexane film which is 120 nm thick at one end of these grooves, and 10 nm at the other, is deposited under a diffusion mask. Reactive etching of the patterned sample transfers a gradient of groove pitch and groove depth into the silicon substrate. A silicon master with a gradient of groove depth spanning more than two orders of magnitude ( less than 10 nm to over 1000 nm) is used to create an injection molding inlay for mass replication of the screening topography. Polycarbonate replicas are molded for use in cell culture studies, and the functionality of the topography as a high-throughput screening platform is investigated. The response of MDCK, h-TERT fibroblasts, and LE2 endothelial cells is examined, in terms of attachment and morphological response to the variation in topographical cues, with the aim of pinpointing the optimal combination of groove pitch and depth to elicit a tailored response from each cell type. When the range of topographical features screened on a single substrate is considered, this new assay represents a significant step forward in the parametric design and analysis of topographical cues at the biomaterial interface.