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While shear emulsification is a well understood industrial process, geometrical confinement in microfluidic systems introduces fascinating complexity, so far prohibiting complete understanding of droplet formation. The size of confined droplets is controlled by the ratio between shear and capillary forces when both are of the same order, in a regime known as jetting, while being surprisingly insensitive to this ratio when shear is orders of magnitude smaller than capillary forces, in a regime known as squeezing. Here, we reveal that further reduction of—already negligibly small—shear unexpectedly re-introduces the dependence of droplet size on shear/capillary-force ratio. For the first time we formally account for the flow around forming droplets, to predict and discover experimentally an additional regime—leaking. Our model predicts droplet size and characterizes the transitions from leaking into squeezing and from squeezing into jetting, unifying the description for confined droplet generation, and offering a practical guide for applications. T-junctions are a tool for droplet generation; they are well-described by models that distinguish for squeezing and jetting regimes for different capillary numbers. By considering the usually neglected corner flow, the authors identify an additional leaking regime for very low capillary numbers.

Since antibiotic resistance is a major threat to global health, recent observations that the traditional test of minimum inhibitory concentration (MIC) is not informative enough to guide effective antibiotic treatment are alarming. Bacterial heteroresistance, in which seemingly susceptible isogenic bacterial populations contain resistant sub-populations, underlies much of this challenge. To close this gap, here we developed a droplet-based digital MIC screen that constitutes a practical analytical platform for quantifying the single-cell distribution of phenotypic responses to antibiotics, as well as for measuring inoculum effect with high accuracy. We found that antibiotic efficacy is determined by the amount of antibiotic used per bacterial colony forming unit (CFU), not by the absolute antibiotic concentration, as shown by the treatment of beta-lactamase-carrying Escherichia coli with cefotaxime. We also noted that cells exhibited a pronounced clustering phenotype when exposed to near-inhibitory amounts of cefotaxime. Overall, our method facilitates research into the interplay between heteroresistance and antibiotic efficacy, as well as research into the origin and stimulation of heterogeneity by exposure to antibiotics. Due to the absolute bacteria quantification in this digital assay, our method provides a platform for developing reference MIC assays that are robust against inoculum-density variations.

Bacterial cells in general present different phenotypes even within the monoclonal populations. Some phenotypes may offer selective advantages under antibiotic stress. In clinical practice such advantage may lead to selection of resistant phenotypes and to treatment failure. Improving treatment regimens thus requires thorough understanding of the underlying cause of developing phenotypic variations within bacterial populations exposed to antibiotics. Here, we use droplet microfluidics to study the response of individual bacterial cells. We investigate how a short-term exposure of ciprofloxacin or streptomycin at the sub-minimum inhibitory concentration levels changes the single-cell distribution of susceptibility on subsequent exposure to either of the antibiotics. We used three different concentrations of pre-exposure for each antibiotic on Escherichia coli MG1655 and followed with re-exposure of the same bacterial cells individually in microfluidic droplets. Experiments with various concentrations of re-exposure on single cells showed that pre-exposures to sub-lethal levels of ciprofloxacin can elevate the individual cell MIC and the level of heteroresistance to ciprofloxacin. Exposure to quarter and half MIC of ciprofloxacin in separate experiments increased the heteroresistance of pre-exposed cells by more than 20-fold on subsequent treatment with ciprofloxacin. Interestingly, the same ciprofloxacin pre-exposure pattern made the cells more fragile and, in some cases, decreased the individual MIC values to streptomycin. The results in our study can be very important in developing new better treatment regimen, addressing the wider problem of antibiotic resistance.

One-dimensional conductive particle assembly holds promise for a variety of practical applications, in particular for a new generation of electronic devices. However, synthesis of such chains with programmable shapes outside a liquid environment has proven difficult. Here we report a route to simply ‘pull’ flexible granular and colloidal chains out of a dispersion by combining field-directed assembly and capillary effects. These chains are automatically stabilized by liquid bridges formed between adjacent particles, without the need for continuous energy input or special particle functionalization. They can further be deposited onto any surface and form desired conductive patterns, potentially applicable to the manufacturing of simple electronic circuits. Various aspects of our route, including the role of particle size and the voltages needed, are studied in detail. Looking towards practical applications, we also present the possibility of two-dimensional writing, rapid solidification of chains and methods to scale up chain production. Conductive colloidal chains are promising for electronics but difficult to synthesize outside of a liquid environment. Here, the authors use field-directed assembly and capillary effects to pull conductive particle chains out of a suspension, which remain held together by flexible liquid bridges even after the external field is turned off.

Droplets of one liquid suspended in a second, immiscible liquid move through a microfluidic device in which a channel splits into two branches that reconnect downstream. The droplets choose a path based on the number of droplets that occupy each branch. The interaction among droplets in the channels results in complex sequences of path selection. The linearity of the flow through the microchannels, however, ensures that the behavior of the system can be reversed. This reversibility makes it possible to encrypt and decrypt signals coded in the intervals between droplets. The encoding/decoding device is a functional microfluidic system that requires droplets to navigate a network in a precise manner without the use of valves, switches, or other means of external control.

The importance of skeletal muscle tissue is undoubted being the controller of several vital functions including respiration and all voluntary locomotion activities. However, its regenerative capability is limited and significant tissue loss often leads to a chronic pathologic condition known as volumetric muscle loss. Here, we propose a biofabrication approach to rapidly restore skeletal muscle mass, 3D histoarchitecture, and functionality. By recapitulating muscle anisotropic organization at the microscale level, we demonstrate to efficiently guide cell differentiation and myobundle formation both in vitro and in vivo . Of note, upon implantation, the biofabricated myo‐substitutes support the formation of new blood vessels and neuromuscular junctions—pivotal aspects for cell survival and muscle contractile functionalities—together with an advanced muscle mass and force recovery. Altogether, these data represent a solid base for further testing the myo‐substitutes in large animal size and a promising platform to be eventually translated into clinical scenarios. Synopsis The regenerative capability of skeletal muscle tissue is limited and significant tissue loss often leads to a chronic pathologic condition known as volumetric muscle loss. By exploiting the potentials of our biofabrication approach, one can manufacture advanced cell‐laden myo‐substitutes that ultimately may restore the functionalities of severely damaged skeletal muscles in vivo . Biofabricated myo‐substitutes may represent a valid candidate for volumetric muscle loss treatment. Upon implantation, biofabricated myo‐substitutes support the formation of new blood vessels and neuromuscular junctions together with an advanced muscle mass and force recovery. The employed biofabrication approach is compatible with human primary stem cells ‐ namely pericytes. Graphical Abstract The regenerative capability of skeletal muscle tissue is limited and significant tissue loss often leads to a chronic pathologic condition known as volumetric muscle loss. By exploiting the potentials of our biofabrication approach, one can manufacture advanced cell‐laden myo‐substitutes that ultimately may restore the functionalities of severely damaged skeletal muscles in vivo .

It is difficult to harness the power generated by biological motors to carry out mechanical work in systems outside the cell. Efforts to capture the mechanical energy of nanomotors ex vivo require in vitro reconstitution of motor proteins and, often, protein engineering. This study presents a method for harnessing the power produced by biological motors that uses intact cells. The unicellular, biflagellated algae Chlamydomonas reinhardtii serve as \"microoxen.\" This method uses surface chemistry to attach loads (1- to 6-μm-diameter polystyrene beads) to cells, phototaxis to steer swimming cells, and photochemistry to release loads. These motile microorganisms can transport microscale loads (3-μm-diameter beads) at velocities of ≈100-200 μ m· sec-1and over distances as large as 20 cm.

Emulsion—a liquid dispersed in another liquid—is in many respects very similar to granular matter. In the early 2000s a new technology—droplet microfluidics—began emerging from the wider field of microfluidics. Droplet microfluidics was quickly established as a discipline of science and engineering and has been used for the generation of highly uniform emulsions. The last few years have brought significant advances to the field, directed towards a wide range of applications in material sciences—from synthesis of nanoparticles in droplets to assembling complex droplets and using droplets as templates for ordered materials, with applications in food, cosmetic and diagnostic industries. Droplet microfluidic platforms are also successfully used as analytical tools in molecular biology and biochemistry, in e.g. high-throughput screening, digital assays, encapsulation of single cells, sequencing technologies, and in point-of-care diagnostic applications. This article provides an accessible overview of the physical phenomena observed in multiphase flow at the microscale and the techniques in droplet microfluidics systems. We also present the most interesting applications and potential further directions of research in this fascinating young field of science and engineering.

Bacterial populations can display different susceptibilities to antibiotics among individual cells, even though they originate from the same parent cell. This variability can lead to treatment failure and the emergence of resistant bacteria. Understanding the factors influencing this variability is crucial for developing effective antibiotic treatments. The underlying cause of variation in susceptibility distribution within bacterial populations remains unclear, necessitating the development of new tools for measurement. Here, we use a droplet microfluidic single-cell antibiotic susceptibility assay and focus on antibiotic resistance conveyed by the TEM β-lactamases family. We investigate how the catalytic activity of β-lactamase, and the genetic characteristics of the host strains affect the susceptibility distribution within bacterial populations at the single-cell level. For this purpose, we selected TEM-1 with the least catalytic activity against cefotaxime, followed by its two variants, R164S and G238S, exhibiting moderate and significant catalytic activity, respectively. The results showed that increasing the catalytic activity causes an increase in the population’s mean level of antibiotic resistance. While the type of β-lactamase influences the susceptibility distribution of the strains, this effect is independent of the catalytic activity of the strains. Besides, the genetic characteristics of the strains receiving the β-lactam resistance gene is an important factor that plays a role in the distribution of susceptibility.

We report the results of a comparative study of microfluidic emulsification of liquids with different viscosities. Depending on the properties of the fluids and their rates of flow, emulsification occurred in the dripping and jetting regimes. We studied the characteristic features and typical dependence of the size and of the size distribution of droplets in each regime. For each liquid, we identified a range of hydrodynamic conditions promoting generation of highly monodisperse droplets. Viscosity played an important role in emulsification: highly viscous liquids were emulsified into larger droplets with lower polydispersity. Although it was not possible to provide a unified scaling for the volumes of the droplets, our results suggest that the break-up dynamics of the lower viscosity fluids resembles the rate-of-flow-controlled break-up, as reported earlier for the formation of bubbles in flow-focusing geometries [Garstecki P, Stone HA, Whitesides GM (2005) Phys Rev Lett 94:164501]. The results of this study can be helpful for a rationalized selection of liquids for the controlled formation of droplets with a predetermined size and with a narrow distribution of sizes.