Fluidic system with movable layers for all-in-one assay of cell-free DNA in blood

Detection of circulating cell-free DNA (cfDNA) requires multiple processes, including plasma separation, cfDNA isolation, and high-sensitivity detection. However, implementing such processes in a single system in a simple and effective manner is nontrivial. Here, we present a fluidic system for the isolation and detection of cfDNA from whole blood performed in a fully automated and effective manner. The system utilizes vertically mobile top and rotational bottom parts with multiple disposable chambers for sequential processes. The key components for plasma filtration are a sedimentation chamber that enables the dextran-enhanced sedimentation of red blood cells and a plunger chamber with a porous filter and a check valve. cfDNA in the plasma is captured by silica-coated magnetic beads in the plunger chamber, washed via the motion of the top and bottom parts, and then detected by real-time on-chip polymerase chain reaction. We vary several parameters, including the plunger speed, dextran concentration, and hematocrit concentration, to optimize the time and efficiency of the assay. The total assay time is 1.5 h with a detection limit of 42 ag/mL for a sample volume of 1.5 mL. Plasma and cfDNA recoveries are 53% and 28%, respectively. Our system is a useful tool for all-in-one assays in liquid biopsies of cancers and other diseases detectable by cfDNA.