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Monitoring microbial reactions in highly opaque or autofluorescent environments like soils, seawater, and wastewater remains challenging. To develop a simple approach for observing post-translational reactions within microbes situated in environmental matrices, we designed a methyl halide transferase (MHT) fragment complementation assay that reports by synthesizing an indicator gas. We show that backbone fission within regions of high sequence variability in the Rossmann domain yields split MHT (sMHT) AND gates whose fragments cooperatively associate to synthesize CH 3 Br. Additionally, we identify a sMHT whose fragments require fusion to pairs of interacting partner proteins for maximal activity. We also show that sMHT fragments fused to FKBP12 and the FKBP-rapamycin binding domain of mTOR display significantly enhanced CH 3 Br production in the presence of rapamycin. This gas production is reversed in the presence of the competitive inhibitor of FKBP12/FKPB dimerization, indicating that sMHT is a reversible reporter of post-translational reactions. This sMHT represents the first genetic AND gate that reports on protein-protein interactions via an indicator gas. Because indicator gases can be measured in the headspaces of complex environmental samples, this assay should be useful for monitoring the dynamics of diverse molecular interactions within microbes situated in hard-to-image marine and terrestrial matrices.

The vasculature is an essential component of the circulatory system that plays a vital role in the development, homeostasis, and disease of various organs in the human body. The ability to emulate the architecture and transport function of blood vessels in the integrated context of their associated organs represents an important requirement for studying a wide range of physiological processes. Traditional in vitro models of the vasculature, however, largely fail to offer such capabilities. Here we combine microfluidic three-dimensional (3D) cell culture with the principle of vasculogenic self-assembly to engineer perfusable 3D microvascular beds in vitro . Our system is created in a micropatterned hydrogel construct housed in an elastomeric microdevice that enables coculture of primary human vascular endothelial cells and fibroblasts to achieve de novo formation, anastomosis, and controlled perfusion of 3D vascular networks. An open-top chamber design adopted in this hybrid platform also makes it possible to integrate the microengineered 3D vasculature with other cell types to recapitulate organ-specific cellular heterogeneity and structural organization of vascularized human tissues. Using these capabilities, we developed stem cell-derived microphysiological models of vascularized human adipose tissue and the blood-retinal barrier. Our approach was also leveraged to construct a 3D organotypic model of vascularized human lung adenocarcinoma as a high-content drug screening platform to simulate intravascular delivery, tumor-killing effects, and vascular toxicity of a clinical chemotherapeutic agent. Furthermore, we demonstrated the potential of our platform for applications in nanomedicine by creating microengineered models of vascular inflammation to evaluate a nanoengineered drug delivery system based on active targeting liposomal nanocarriers. These results represent a significant improvement in our ability to model the complexity of native human tissues and may provide a basis for developing predictive preclinical models for biopharmaceutical applications.

Diverse biological processes in the body rely on the ability of cells to exert contractile forces on their extracellular matrix (ECM). In three-dimensional (3D) cell culture, however, this intrinsic cellular property can cause unregulated contraction of ECM hydrogel scaffolds, leading to a loss of surface anchorage and the resultant structural failure of in vitro tissue constructs. Despite advances in the 3D culture technology, this issue remains a significant challenge in the development and long-term maintenance of physiological 3D in vitro models. Here, we present a simple yet highly effective and accessible solution to this problem. We leveraged a single-step surface functionalization technique based on polydopamine to drastically increase the strength of adhesion between hydrogel scaffolds and cell culture substrates. Our method is compatible with different types of ECM and polymeric surfaces and also permits prolonged shelf storage of functionalized culture substrates. The proof-of-principle of this technique was demonstrated by the stable long-term (1 month) 3D culture of human lung fibroblasts. Furthermore, we showed the robustness and advanced application of the method by constructing a dynamic cell stretching system and performing over 100 000 cycles of mechanical loading on 3D multicellular constructs for visualization and quantitative analysis of stretch-induced tissue alignment. Finally, we demonstrated the potential of our technique for the development of microphysiological in vitro models by establishing microfluidic 3D co-culture of vascular endothelial cells and fibroblasts to engineer self-assembled, perfusable 3D microvascular beds.

Century-old Langmuir monolayer deposition still represents the most convenient approach to the production of monolayers of colloidal nanoparticles on solid substrates for practical biological and chemical-sensing applications. However, this approach simply yields arbitrarily shaped large monolayers on a flat surface and is strongly limited by substrate topography and interfacial energy. Here, we describe a generalized and facile method of rapidly producing uniform monolayers of various colloidal nanoparticles on arbitrary solid substrates by using an ordinary capillary tube. Our method is based on an interesting finding of inversion phenomenon of a nanoparticle-laden air-water interface by flowing through a capillary tube in a manner that prevents the particles from adhesion to the capillary sidewall, thereby presenting the nanoparticles face-first at the tube's opposite end for direct and one-step deposition onto a substrate. We show that our method not only allows the placement of a nanoparticle monolayer at target locations of solid substrates regardless of their surface geometry and adhesion but also enables the production of monolayers containing nanoparticles with different size, shape, surface charge, and composition. To explore the potential of our approach, we demonstrate the facile integration of gold nanoparticle monolayers into microfluidic devices for the real-time monitoring of molecular Raman signals under dynamic flow conditions. Moreover, we successfully extend the use of our method to developing on-demand Raman sensors that can be built directly on the surface of consumer products for practical chemical sensing and fingerprinting. Specifically, we achieve both the pinpoint deposition of gold nanoparticle monolayers and sensitive molecular detection from the deposited region on clothing fabric for the detection of illegal drug substances, a single grain of rice and an orange for pesticide monitoring, and a $100 bill as a potential anti-counterfeit measure, respectively. We believe that our method will provide unique opportunities to expand the utility of colloidal nanoparticles and to greatly improve the accessibility of nanoparticle-based sensing technologies.

For low-cost and facile fabrication of innovative nanoscale devices with outstanding functionality and performance, it is critical to develop more practical patterning solutions that are applicable to a wide range of materials and feature sizes while minimizing detrimental effects by processing conditions. In this study, we report that area-selective sub-10 nm pattern formation can be realized by temperature-controlled spin-casting of block copolymers (BCPs) combined with submicron-scale-patterned chemical surfaces. Compared to conventional room-temperature spin-casting, the low temperature ( e.g., -5 °C) casting of the BCP solution on the patterned self-assembled monolayer achieved substantially improved area selectivity and uniformity, which can be explained by optimized solvent evaporation kinetics during the last stage of film formation. Moreover, the application of cold spin-casting can also provide high-yield in situ patterning of light-emitting CdSe/ZnS quantum dot thin films, indicating that this temperature-optimized spin-casting strategy would be highly effective for tailored patterning of diverse organic and hybrid materials in solution phase.

Detection and control of air quality are major concerns in recent years for environmental monitoring and healthcare. In this work, we developed an integrated sensor architecture comprised of nanostructured composite sensing layers and a flexible heating substrate for portable and real-time detection of nitrogen dioxide (NO 2 ). As sensing layers, PdO-infiltrated Co 3 O 4 hollow nanocubes (PdO-Co 3 O 4 HNCs) were prepared by calcination of Pd-embedded Co-based metal-organic framework polyhedron particles. Single-walled carbon nanotubes (SWCNTs) were functionalized with PdO-Co 3 O 4 HNCs to control conductivity of sensing layers. As a flexible heating substrate, the Ni mesh electrode covered with a 40 nm thick Au layer (i.e., Ni(core)/Au(shell) mesh) was embedded in a colorless polyimide (cPI) film. As a result, SWCNT-functionalized PdO-Co 3 O 4 HNCs sensor exhibited improved NO 2 detection property at 100 °C, with high sensitivity (S) of 44.11% at 20 ppm and a low detection limit of 1 ppm. The accelerated reaction and recovery kinetics toward NO 2 of SWCNT-functionalized PdO-Co 3 O 4 HNCs were achieved by generating heat on the Ni(core)/Au(shell) mesh-embedded cPI substrate. The SWCNT-functionalized porous metal oxide sensing layers integrated on the mechanically stable Ni(core)/Au(shell) mesh heating substrate can be envisioned as an essential sensing platform for realization of low-temperature operation wearable chemical sensor.

We describe a bioinspired microfluidic system that resembles pulmonary airways and enables on-chip generation of airway occluding liquid plugs from a stratified air-liquid two-phase flow. User-defined changes in the air stream pressure facilitated by mechanical components and tuning the wettability of the microchannels enable generation of well-defined liquid plugs. Significant differences are observed in liquid plug generation and propagation when surfactant is added to the buffer. The plug flow patterns suggest a protective role of surfactant for airway epithelial cells against pathological flow-induced mechanical stresses. We discuss the implications of the findings for clinical settings. This approach and the described platform will enable systematic investigation of the effect of different degrees of fluid mechanical stresses on lung injury at the cellular level and administration of exogenous therapeutic surfactants.

The integration of semiporous membranes into poly(dimethylsiloxane) (PDMS) microfluidic devices is useful for mass transport control. Several methods such as plasma oxidation and manual application of PDMS prepolymer exist to sandwich such membranes into simple channel structures, but these methods are difficult to implement with reliable sealing and no leakage or clogging for devices with intricate channel features. This paper describes a simple but robust strategy to bond semiporous polyester and polycarbonate membranes between layers of PDMS microchannel structures effectively without channel clogging. A thin layer of PDMS prepolymer, spin-coated on a glass slide, is transferred to PDMS substrates with channel features as well as to the edges of the semiporous membrane by stamping. This thin PDMS prepolymer serves as "mortar" to strongly bond the two PDMS layers and seal off the crevices generated from the thickness of the membranes. This bonding method enabled the fabrication of an 8x12 criss-crossing microfluidic channel array with 96 combinations of fluid interactions. The capability of this device for bioanalysis was demonstrated by measuring responses of cells to different color fluorescent reagents.

This paper describes a simple microfluidic sorting system that can perform size profiling and continuous mass-dependent separation of particles through combined use of gravity (1 g) and hydrodynamic flows capable of rapidly amplifying sedimentation-based separation between particles. Operation of the device relies on two microfluidic transport processes: (i) initial hydrodynamic focusing of particles in a microchannel oriented parallel to gravity and (ii) subsequent sample separation where positional difference between particles with different mass generated by sedimentation is further amplified by hydrodynamic flows whose streamlines gradually widen out due to the geometry of a widening microchannel oriented perpendicular to gravity. The microfluidic sorting device was fabricated in poly(dimethylsiloxane), and hydrodynamic flows in microchannels were driven by gravity without using external pumps. We conducted theoretical and experimental studies on fluid dynamic characteristics of laminar flows in widening microchannels and hydrodynamic amplification of particle separation. Direct trajectory monitoring, collection, and post-analysis of separated particles were performed using polystyrene microbeads with different sizes to demonstrate rapid (<1 min) and high-purity (>99.9%) separation. Finally, we demonstrated biomedical applications of our system by isolating small-sized (diameter <6 microm) perfluorocarbon liquid droplets from polydisperse droplet emulsions, which is crucial in preparing contrast agents for safe, reliable ultrasound medical imaging, tracers for magnetic resonance imaging, or transpulmonary droplets used in ultrasound-based occlusion therapy for cancer treatment. Our method enables straightforward, rapid, real-time size monitoring and continuous separation of particles in simple stand-alone microfabricated devices without the need for bulky and complex external power sources. We believe that this system will provide a useful tool to separate colloids and particles for various analytical and preparative applications and may hold potential for separation of cells or development of diagnostic tools requiring point-of-care sample preparation or testing.

This work is the first demonstration of electrical modulation of surface energy to reversibly switch dynamic high-speed gas-liquid two-phase microfluidic flow patterns. Manipulation of dynamic two-phase systems with continuous high-speed flows is complex and interesting due to the multiple types of forces that need to be considered. Here, distinct stable flow patterns are formed through a multipronged approach: both surface tension forces generated by surface chemistry modulation as well as viscous and inertial forces produced by fluid flows are employed. The novel fluidic actuation mechanism provides insights into better understanding microscale two-phase flow dynamics and offers new opportunities for the development of two-phase biochemical microsystems that are mechanically simple and operational at high speeds.