Han Jung Y, Wiederoder Michael, DeVoe Don L
1Department of Mechanical Engineering, University of Maryland, College Park, MD 20742 USA.
2Department of Chemical and Biomolecular Engineering, University of Maryland, College Park, MD 20742 USA.
Microsyst Nanoeng. 2019 Jun 17;5:30. doi: 10.1038/s41378-019-0063-4. eCollection 2019.
Rapid and efficient isolation of bacteria from complex biological matrices is necessary for effective pathogen identification in emerging single-cell diagnostics. Here, we demonstrate the isolation of intact and viable bacteria from whole blood through the selective lysis of blood cells during flow through a porous silica monolith. Efficient mechanical hemolysis is achieved while providing passage of intact and viable bacteria through the monoliths, allowing size-based isolation of bacteria to be performed following selective lysis. A process for synthesizing large quantities of discrete capillary-bound monolith elements and millimeter-scale monolith bricks is described, together with the seamless integration of individual monoliths into microfluidic chips. The impact of monolith morphology, geometry, and flow conditions on cell lysis is explored, and flow regimes are identified wherein robust selective blood cell lysis and intact bacteria passage are achieved for multiple gram-negative and gram-positive bacteria. The technique is shown to enable rapid sample preparation and bacteria analysis by single-cell Raman spectrometry. The selective lysis technique presents a unique sample preparation step supporting rapid and culture-free analysis of bacteria for the point of care.
在新兴的单细胞诊断中，要有效鉴定病原体，就需要从复杂生物基质中快速高效地分离细菌。在此，我们展示了在流经多孔硅胶整体柱时通过选择性裂解血细胞从全血中分离完整且有活力的细菌。在使完整且有活力的细菌通过整体柱的同时实现了高效的机械溶血，从而在选择性裂解后能够基于大小对细菌进行分离。描述了一种合成大量离散的毛细管连接整体柱元件和毫米级整体柱砖的方法，以及将单个整体柱无缝集成到微流控芯片中的方法。探讨了整体柱形态、几何形状和流动条件对细胞裂解的影响，并确定了多种革兰氏阴性菌和革兰氏阳性菌能够实现强力选择性血细胞裂解和完整细菌通过的流动状态。结果表明，该技术能够通过单细胞拉曼光谱法实现快速样品制备和细菌分析。这种选择性裂解技术提供了一个独特的样品制备步骤，支持即时护理时对细菌进行快速且无需培养的分析。
