Full-Combinatorial Concentration Gradient Array with 3D Micro/Nanofluidics for Antibiotic Susceptibility Testing.

Recent advancements in micro/nanofluidics have facilitated on-chip microscopy of cellular responses in a high-throughput and controlled microenvironment with the desired physicochemical properties. Despite its potential benefits to combination drug discovery, generating a complete combinatorial set of concentration gradients for multiple reagents in an array format remains challenging. The main reason is limited layouts of conventional micro/nanofluidic systems based on two-dimensional channel networks. In this paper, we present a device with three-dimensional (3D) interconnection of micro/nanochannels capable of generating a complete combinatorial set of concentration gradients for two reagents. The device was readily fabricated by laminating a pair of multilayered monolithic films containing a Christmas tree-like mixer, a cell culture chamber array, and through-holes, all within each single film. We assessed the reliable generation of a full-combinatorial concentration gradient array and validated it by using numerical analysis. We applied the proposed device to test the antibiotic susceptibility of bacterial cells in a convenient one-step manner. Furthermore, we explored the potential of the device to accommodate the arrayed complete combinatorial set for two or more drugs, while extending the capabilities of our laminated object manufacturing method for realizing 3D micro/nanofluidic systems.

Pervaporation-assisted in situ formation of nanoporous microchannels with various material and structural properties.

Nanoporous structures are crucial for developing mixed-scale micro-/nanofluidic devices because they facilitate the manipulation of molecule transport along the microfluidic channel networks. Particularly, self-assembled particles have been used for fabricating various nanoporous membranes. However, previous self-assembly mechanisms relied on the material and structural homogeneities of the nanopores. Here, we present a pervaporation-assisted in situ fabrication method that integrates nanoporous membrane structures into microfluidic devices. The microfluidic devices contain a control-channel layer at the top, which induces local and addressable pervaporation, and the main-channel layer, which is present at the bottom with pre-designated locations for nanoporous microchannels; the layers are separated using a gas-permeable film. The target particle suspensions are loaded into the main channels, and their pervaporation is controlled through the gas-permeable film, which successfully assembles the particles at the pre-designated locations. This method yields nanoporous microchannels with various material and structural properties by fabricating heterogeneous nanopore arrays/junctions in series and other diverse structures along the microchannels. We validate the basic working principle of microfluidic devices containing nanoporous microchannels. Furthermore, we theoretically analyze the fundamental experimental results, which suggest the remarkable potential of our strategy to fabricate nanopore networks without using conventional nanofabrication methods.