<div>A micro- and nano-fluidic device stacked with magnetic beads is developed to efficiently trap, concentrate, and retrieve Escherichia coli (E. coli) from bacteria suspension</div><div>and pig plasma. The small voids between the magnetic beads are used to physically isolate the bacteria in the device. We use computational fluid dynamics (CFD), 3D</div><div>tomography technology, and machine learning to probe and explain the bead stacking in a small 3D space with various flow rates. A combination of beads with different sizes is utilized to achieve a high capture efficiency of ~86% with a flow rate of 50 μL/min. Leveraging the high deformability of this device, the E. coli sample is retrieved from the designated bacteria suspension by applying a higher flow rate, followed by rapid magnetic separation. This unique function is also utilized to concentrate E. coli from the original bacteria suspension. An on-chip concentration</div><div>factor of ~11× is achieved by inputting 1,300 μL of the E. coli sample and then concentrating it in 100 μL buffer.</div><div>Importantly, this multiplexed, miniaturized, inexpensive, and transparent device is easy to fabricate and operate, making it ideal for pathogen separation in both laboratory and pointof- care (POC) settings.</div>