Department of Chemistry, University of Kansas, Lawrence, KS 66045, USA.
Lab Chip. 2018 Dec 4;18(24):3830-3839. doi: 10.1039/c8lc00927a.
Rapid, sensitive analysis of protein biomarkers is of tremendous biological and clinical significance. Immunoassays are workhorse tools for protein analysis and have been under continuous investigation to develop new methods and to improve the analytical performance. Herein we report a pneumatically gated microfluidic communicating vessel (μCOVE) chip for rapid and sensitive immunomagnetic ELISA. A distinct feature of our device is that it employs the communicating vessel principle as a simple means to generate a fast transient hydrodynamic flow to enable effective flow washing without the need for excessive incubation, which greatly simplifies and expedites the assay workflow, compared to conventional microfluidic flow-based immunoassays. Stationary multi-phase microfluidic techniques have been developed for fast bead washing. However, they have some limitations, such as the need for careful control of interfacial properties, large bead quantity required for reliable interphase bead transport, and relatively high bead loss during surface tension-gated traverse. Our single-phase μCOVE chip can overcome such limitations and facilitate the manipulation of magnetic beads to streamline the assay workflow. We showed that the μCOVE device affords highly sensitive quantification of the CEA and EGFR proteins with a LOD down to the sub-picogram per mL level. Direct detection of the EGFR in the crude A431 cell lysate was also demonstrated to further validate the ability of our device for rapid and quantitative analysis of complex biological samples. Overall, our work presents a unique platform that combines the merits of the stationary multi-phase systems and the flow-based microfluidics. This novel immunoassay microsystem has promising potential for a broad range of biological and clinical applications, owing to its simplicity and high performance.
快速、灵敏地分析蛋白质生物标志物具有巨大的生物学和临床意义。免疫测定法是蛋白质分析的主要工具，一直在不断研究开发新方法和提高分析性能。在此，我们报告了一种气动门控微流控连通容器（μCOVE）芯片，用于快速灵敏的免疫磁酶联免疫吸附测定法（ELISA）。我们的装置的一个显著特点是它采用连通容器原理作为一种简单的方法来产生快速瞬态流体流动，从而能够有效地进行流动洗涤，而无需过度孵育，与传统的微流控基于流动的免疫测定法相比，大大简化和加快了测定工作流程。固定多相微流控技术已被开发用于快速珠洗涤。然而，它们存在一些限制，例如需要仔细控制界面特性、可靠的相间珠传输所需的大量珠以及在表面张力门控横越过程中相对较高的珠损失。我们的单相μCOVE 芯片可以克服这些限制，并方便磁珠的操作，简化测定工作流程。我们表明，μCOVE 装置能够高度灵敏地定量检测 CEA 和 EGFR 蛋白，LOD 低至亚皮克每毫升水平。还直接检测了粗 A431 细胞裂解物中的 EGFR，进一步验证了我们的设备用于快速定量分析复杂生物样品的能力。总的来说，我们的工作提出了一个独特的平台，结合了固定多相系统和基于流动的微流控的优点。由于其简单性和高性能，这种新型免疫分析微系统具有广泛的生物学和临床应用的巨大潜力。
