界面に着目した高機能バイオマーカー検出マイクロ流路システムに関する研究 Surface-based Microfluidic Systems for Enhanced Biomarker Detection

The 21st century has seen a surge in the development of point-of-care (POC) testing systems integrated with microfluidic bioassay devices, for portable, fast, and user-friendly disease diagnostics. These systems detect diagnostic biomarkers from a small quantity of the patient’s blood plasma, by exploiting their innate nature to bind to specific receptor molecules. A microfluidic bioassay device is considered to be of “high efficiency” when low biomarker concentrations (1 pM–1 nM) can be detected within a few minutes. This thesis explores the collective influence of surface chemistry, biomarker transport and biomolecular reactions at the microscale, to propose design principles for the development of rapid, sensitive and user-friendly fluorescence-based POC systems. First, we exploit radio-frequency air plasma to covalently tether receptor proteins within polymethyl methacrylate microfluidic bioassay devices, at high-throughput. Next, these devices are integrated with a palm-sized modular Fluid Handling Device that allows precise mixing, filtration, and delivery of fluids, for subsequent detection of Chlamydia trachomatis specific antibodies, with a limit of detection (LoD) of 7 nM within 15 mins, serving as a “proof-of-concept” POC testing device. Next, biomarker transport-dependent kinetic enhancements in microfluidic bioassay systems are investigated using novel 3D glass devices, where real-time binding events between varying concentrations of fluorescently-labelled receptor and ligand antibodies are analyzed. Combing experimental measurements with scaling analysis, two key control dimensionless parameters are proposed to achieve “rapid” and “sensitive” ligand detection: a local Peclet number P eδ that characterizes the balance between local convection and diffusion-driven transport of ligands; and a kinetic Damkohler number (Dakinetic) that characterizes the balance between the rates of receptor–ligand binding and convection-driven ligand replenishment. We observe that homogeneous ligand binding can be achieved by decreasing the depletion layer thickness (< 5 µm) with enhanced local convection at P eδ >> 105. At Dakinetic << 10−2, we demonstrate that enhanced convective ligand replenishment leads to quenching of transport limitations, thereby enhancing the speed of ligand detection at the molecular limit. Finally, we validate that rapid (detection time of 10 mins) and sensitive (LoD of 11.63 pM) ligand detection can be achieved in microfluidic systems with P eδ >> 107 for Dakinetic << 10−2. With prior knowledge of the kinetic constants, these design principles can be applied to various biomolecular systems, paving way to creating highly efficient POC testing systems in the near future.