Department of Chemistry, University of Virginia, McCormick Road, P.O. Box 400319, Charlottesville, Virginia 22904, USA.
Lab Chip. 2011 May 7;11(9):1603-11. doi: 10.1039/c0lc00597e. Epub 2011 Mar 4.
This work describes the performance of poly(methyl methacrylate) (PMMA) microfluidic DNA purification devices with embedded microfabricated posts, functionalized with chitosan. PMMA is attractive as a substrate for creating high surface area (SA) posts for DNA capture because X-ray lithography can be exploited for extremely reproducible fabrication of high SA structures. However, this advantage is offset by the delicate nature of the posts when attempting bonding to create a closed system, and by the challenge of functionalizing the PMMA surface with a group that invokes DNA binding. Methods are described for covalent functionalization of the post surfaces with chitosan that binds DNA in a pH-dependent manner, as well as for bonding methods that avoid damaging the underlying post structure. A number of geometric posts designs are explored, with the goal of identifying post structures that provide the requisite surface area without a concurrent rise in fluidic resistance that promotes device failure. Initial proof-of-principle is shown by recovery of prepurified human genomic DNA (hgDNA), with real-world utility illustrated by purifying hgDNA from whole blood and demonstrating it to be PCR-amplifiable.
这项工作描述了具有嵌入式微加工柱的聚甲基丙烯酸甲酯（PMMA）微流控 DNA 纯化设备的性能，这些柱子用壳聚糖功能化。PMMA 作为用于创建用于 DNA 捕获的高表面积 (SA) 柱子的基底具有吸引力，因为 X 射线光刻可以用于非常可重复地制造高 SA 结构。然而，当试图进行键合以创建封闭系统时，这种优势被柱子的脆弱性质抵消了，并且在 PMMA 表面上用引发 DNA 结合的基团进行功能化也具有挑战性。描述了用于通过壳聚糖共价功能化柱表面的方法，壳聚糖以 pH 依赖性的方式结合 DNA，以及避免损坏底层柱结构的键合方法。探索了多种几何柱设计，目的是确定提供必需表面积的柱结构，而不会同时增加促进器件失效的流体阻力。通过从预纯化的人类基因组 DNA（hgDNA）中回收来证明初始原理，通过从全血中纯化 hgDNA 并证明其可进行 PCR 扩增来展示实际应用。
