This study reports an advanced approach to effectively generate hollow fibers in a triple-flow polydimethylsiloxane (PDMS) microfluidic device based on the gelation of alginate induced with CaCl₂ inside a coaxial flow system. Two PDMS replicas with a semi-cylindrical microchannel were assembled to obtain a complete microchannel with a circular cross-section, which allowed the formation of mild and continuous coaxial flows for the fabrication of hollow fibers without employing complex glass microcapillaries. Mineral oil was introduced into the central flow to serve as an inert space inside the Ca–alginate wall. This was used to maintain the consistent formation of the hollow core of the microfiber and to easily transport fluid through the lumen structure in subsequent applications. The hollow fibers exhibited characteristics such as flexibility while showing robust mechanical strength, high permeability, and biocompatibility, and were used as scaffolds for the attachment and proliferation of human umbilical vein endothelial cells (HUVECs) to mimic a blood vessel. The fully covered HUVEC fibers were further integrated into a neurovascular system and co-cultured with astrocytes forming an on-chip blood brain barrier (BBB) platform. The use of this neurovascular model for drug testing will pave the way for developing or synthesizing a new drug that can cross the BBB in the human brain.

Influenza A viruses are often present in environmental and clinical samples at concentrations below the limit of detection (LOD) of molecular diagnostics. Here we report an integrated microfluidic preconcentration and nucleic amplification system (μFPNAS) which enables both preconcentration of influenza A virus H1N1 (H1N1) and amplification of its viral RNA, thereby lowering LOD for H1N1. H1N1 virus particles were first magnetically preconcentrated using magnetic nanoparticles conjugated with an antibody specific for the virus. Their isolated RNA was amplified to cDNA through thermocycling in a trapezoidal chamber of the μFPNAS. A detection limit as low as 100 TCID50 (50% tissue culture infective dose) in saliva can be obtained within 2 hours. These results suggest that the LOD of molecular diagnostics for virus can be lowered by systematically combining immunomagnetic separation and reverse transcriptase-polymerase chain reaction (RT-PCR) in one microfluidic device.

Here, we introduce a simple and fast method for bonding a poly(dimethylsiloxane) (PDMS) silicone elastomer to different plastics. In this technique, surface modification and subsequent bonding processes are performed at room temperature. Furthermore, only one chemical is needed, and no surface oxidation step is necessary prior to bonding. This bonding method is particularly suitable for encapsulating biomolecules that are sensitive to external stimuli, such as heat or plasma treatment, and for embedding fracturable materials prior to the bonding step. Microchannel-fabricated PDMS was first oxidized by plasma treatment and reacted with aminosilane by forming strong siloxane bonds (Si–O–Si) at room temperature. Without the surface oxidation of the amine-terminated PDMS and plastic, the two heterogeneous substrates were brought into intimate physical contact and left at room temperature. Subsequently, aminolysis occurred, leading to the generation of a permanent seal via the formation of robust urethane bonds after only 5 min of assembling. Using this method, large-area (10 × 10 cm) bonding was successfully realized. The surface was characterized by contact angle measurements and X-ray photoelectron spectroscopy (XPS) analyses, and the bonding strength was analyzed by performing peel, delamination, leak, and burst tests. The bond strength of the PDMS–polycarbonate (PC) assembly was approximately 409 ± 6.6 kPa, and the assembly withstood the injection of a tremendous amount of liquid with the per-minute injection volume exceeding 2000 times its total internal volume. The thermal stability of the bonded microdevice was confirmed by performing a chamber-type multiplex polymerase chain reaction (PCR) of two major foodborne pathogens – Escherichia coli O157:H7 and Salmonella typhimurium – and assessing the possibility for on-site direct detection of PCR amplicons. This bonding method demonstrated high potential for the stable construction of closed microfluidic systems socketed with biomolecule-immobilized surfaces such as DNA, antibody, enzyme, peptide, and protein microarrays.

The human pathogen Listeria monocytogenes can cope with severe environmental challenges, for which the high molecular weight stressosome complex acts as the sensing hub in a complicated signal transduction pathway. Here, we show the dynamics and functional roles of the stressosome protein RsbR1 and its paralogue, the blue-light receptor RsbL, using photo-activated localization microscopy combined with single-particle tracking and single-molecule displacement mapping and supported by physiological studies. In live cells, RsbR1 is present in multiple states: in protomers with RsbS, large clusters of stressosome complexes, and in connection with the plasma membrane via Prli42. RsbL diffuses freely in the cytoplasm but forms clusters upon exposure to light. The clustering of RsbL is independent of the presence of Prli42. Our work provides a comprehensive view of the spatial organization and intracellular dynamics of the stressosome proteins in L. monocytogenes, which paves the way towards uncovering the stress-sensing mechanism of this signal transduction pathway.

Drug-resistant tuberculosis (DR-TB) kills ~200,000 people every year. A contributing factor is the slow turnaround time (TAT) associated with drug susceptibility diagnostics. The prevailing gold standard for phenotypic drug susceptibility testing (pDST) takes at least two weeks. Here we show that growth-based pDST for slow-growing mycobacteria can be conducted in 12 h. We use Mycobacterium tuberculosis variant bovis Bacillus Calmette-Guérin (BCG) and Mycobacterium smegmatis as the mycobacterial pathogen models and expose them to antibiotics used in (multidrug-resistant) tuberculosis (TB) treatment regimens - i.e., rifampicin (RIF), isoniazid (INH), ethambutol (EMB), linezolid (LZD), streptomycin (STR), bedaquiline (BDQ), and levofloxacin (LFX). The bacterial growth in a microfluidic chip is tracked by time-lapse phase-contrast microscopy. A deep neural network-based segmentation algorithm is used to quantify the growth rate and to determine how the strains responded to drug treatments. Most importantly, a panel of susceptible and resistant M. bovis BCG are tested at critical concentrations for INH, RIF, STR, and LFX. The susceptible strains could be identified in less than 12 h. These findings are comparable to what we expect for pathogenic M. tuberculosis as they share 99.96% genetic identity.