Breadmore Michael C, Wolfe Kelley A, Arcibal Imee G, Leung Wayne K, Dickson Dana, Giordano Braden C, Power Mary E, Ferrance Jerome P, Feldman Sanford H, Norris Pamela M, Landers James P
Department of Chemistry, University of Virginia, Charlottesville, Virginia 22904, USA.
Anal Chem. 2003 Apr 15;75(8):1880-6. doi: 10.1021/ac0204855.
A microchip solid-phase extraction method for purification of DNA from biological samples, such as blood, is demonstrated. Silica beads were packed into glass microchips and the beads immobilized with sol-gel to provide a stable and reproducible solid phase onto which DNA could be adsorbed. Optimization of the DNA loading conditions established a higher DNA recovery at pH 6.1 than 7.6. This lower pH also allowed for the flow rate to be increased, resulting in a decrease in extraction time from 25 min to less than 15 min. Using this procedure, template genomic DNA from human whole blood was purified on the microchip platform with the only sample preparation being mixing of the blood with load buffer prior to loading on the microchip device. Comparison between the microchip SPE (microchipSPE) procedure and a commercial microcentrifuge method showed comparable amounts of PCR-amplifiable DNA could be isolated from cultures of Salmonella typhimurium. The greatest potential of the microchipSPE device was illustrated by purifying DNA from spores from the vaccine strain of Bacillus anthracis, where eventual integration of SPE, PCR, and separation on a single microdevice could potentially enable complete detection of the infectious agent in less than 30 min.
展示了一种用于从生物样品（如血液）中纯化DNA的微芯片固相萃取方法。将硅胶珠填充到玻璃微芯片中，并用溶胶 - 凝胶固定这些珠子，以提供一个稳定且可重复的固相，DNA可吸附在其上。DNA加载条件的优化表明，在pH 6.1时的DNA回收率高于pH 7.6时。较低的pH值还允许提高流速，从而使提取时间从25分钟减少到不到15分钟。使用此程序，在微芯片平台上纯化了来自人全血的模板基因组DNA，唯一的样品制备步骤是在将血液加载到微芯片装置之前将其与加载缓冲液混合。微芯片固相萃取（microchipSPE）程序与商业微量离心法之间的比较表明，从鼠伤寒沙门氏菌培养物中可分离出相当数量的可用于PCR扩增的DNA。通过从炭疽芽孢杆菌疫苗株的孢子中纯化DNA说明了微芯片SPE装置的最大潜力，在单个微装置上最终整合固相萃取、PCR和分离可能能够在不到30分钟内完成对感染因子的完整检测。
