Du K, Cai H, Park M, Wall T A, Stott M A, Alfson K J, Griffiths A, Carrion R, Patterson J L, Hawkins A R, Schmidt H, Mathies R A
Department of Chemistry, University of California at Berkeley, Berkeley, CA 94720, USA.
School of Engineering, University of California Santa Cruz, 1156 High Street, Santa Cruz, CA 95064, USA.
Biosens Bioelectron. 2017 May 15;91:489-496. doi: 10.1016/j.bios.2016.12.071. Epub 2017 Jan 3.
An automated microfluidic sample preparation multiplexer (SPM) has been developed and evaluated for Ebola virus detection. Metered air bubbles controlled by microvalves are used to improve bead-solution mixing thereby enhancing the hybridization of the target Ebola virus RNA with capture probes bound to the beads. The method uses thermally stable 4-formyl benzamide functionalized (4FB) magnetic beads rather than streptavidin coated beads with a high density of capture probes to improve the target capture efficiency. Exploiting an on-chip concentration protocol in the SPM and the single molecule detection capability of the antiresonant reflecting optical waveguide (ARROW) biosensor chip, a detection limit of 0.021pfu/mL for clinical samples is achieved without target amplification. This RNA target capture efficiency is two orders of magnitude higher than previous results using streptavidin beads and the limit of detection (LOD) improves 10×. The wide dynamic range of this technique covers the whole clinically applicable concentration range. In addition, the current sample preparation time is ~1h which is eight times faster than previous work. This multiplexed, miniaturized sample preparation microdevice establishes a key technology that intended to develop next generation point-of-care (POC) detection system.
一种用于埃博拉病毒检测的自动化微流控样品制备多路复用器（SPM）已被开发并评估。由微阀控制的定量气泡用于改善磁珠与溶液的混合，从而增强目标埃博拉病毒RNA与结合在磁珠上的捕获探针的杂交。该方法使用热稳定的4-甲酰基苯甲酰胺功能化（4FB）磁珠，而不是具有高密度捕获探针的链霉亲和素包被磁珠，以提高目标捕获效率。利用SPM中的芯片上浓缩方案和反共振反射光波导（ARROW）生物传感器芯片的单分子检测能力，在无需目标扩增的情况下，临床样品的检测限达到0.021pfu/mL。这种RNA目标捕获效率比以前使用链霉亲和素磁珠的结果高两个数量级，检测限提高了10倍。该技术的宽动态范围覆盖了整个临床适用浓度范围。此外，目前的样品制备时间约为1小时，比以前的工作快八倍。这种多路复用、小型化的样品制备微型设备建立了一项关键技术，旨在开发下一代即时检测（POC）系统。
