Picoliter

Interaction between tumor-associated macrophages and tumor cells is crucial for tumor development, metastasis, and the related immune process. However, the macrophages are highly heterogeneous spanning from anti-tumorigenic to pro-tumorigenic, which needs to be understood at the single-cell level. Herein, a sessile microdroplet system designed for monitoring cellular behavior and analyzing intercellular interaction, demonstrated with macrophage-tumor cell pairs is presented. An automatic procedure based on the inkjet printing method is utilized for the precise pairing and co-encapsulation of heterotypic cells within picoliter droplets...

Traditional methods for the enrichment of microorganisms rely on growth in a selective liquid medium or on an agar plate, followed by tedious characterization. Droplet microfluidic techniques have been recently used to cultivate microorganisms and preserve enriched bacterial taxonomic diversity. However, new methods are needed to select droplets comprising not only growing microorganisms but also those exhibiting specific properties, such as the production of value-added compounds. We describe here a droplet microfluidic screening technique for the functional selection of biosurfactant-producing microorganisms, which are of great interest in the bioremediation and biotechnology industries...

Enzymes are widely used as catalysts in the chemical and pharmaceutical industries. While successful in many situations, they must usually be adapted to operate efficiently under nonnatural conditions. Enzyme engineering allows the creation of novel enzymes that are stable at elevated temperatures or have higher activities and selectivities. Current enzyme engineering techniques require the production and testing of enzyme variant libraries to identify members with desired attributes. Unfortunately, traditional screening methods cannot screen such large mutagenesis libraries in a robust and timely manner...

Inkjet printing is a ubiquitous consumer and industrial process that involves concomitant processes of droplet impact, wetting, evaporation, and imbibement into a substrate as well as consequential substrate rearrangements and remodeling. In this work, we perform a study on the interaction between ink dispersions of different composition on substrates of increasing complexity to disentangle the motion of the liquid from the dynamic response of the substrate. We print three variations of pigmented inks and follow the ensuing dynamics at millisecond and micron time and length scales until complete drying using a multiple scattering technique, laser speckle imaging (LSI)...

The structural determination of natural products (NPs) can be arduous because of sample heterogeneity. This often demands iterative purification processes and characterization of complex molecules that may be available only in miniscule quantities. Microcrystal electron diffraction (microED) has recently shown promise as a method to solve crystal structures of NPs from nanogram quantities of analyte. However, its implementation in NP discovery remains hampered by sample throughput and purity requirements, akin to traditional NP-discovery workflows...

A common challenge in microfluidic cell cultures has to do with analysis of cell function without replacing a significant fraction of the culture volume and disturbing local concentration gradients of signals. To address this challenge, we developed a microfluidic cell culture device with an integrated bioanalysis unit to enable on-chip analysis of picoliter volumes of cell-conditioned media. The culture module consisted of an array of 140 microwells with a diameter of 300 m which were made low-binding to promote organization of cells into 3D spheroids...

Droplet microfluidics enables kHz screening of picoliter samples at a fraction of the cost of other high-throughput approaches. However, generating stable droplets with desired characteristics typically requires labor-intensive empirical optimization of device designs and flow conditions that limit adoption to specialist labs. Here, we compile a comprehensive droplet dataset and use it to train machine learning models capable of accurately predicting device geometries and flow conditions required to generate stable aqueous-in-oil and oil-in-aqueous single and double emulsions from 15 to 250 μm at rates up to 12000 Hz for different fluids commonly used in life sciences...

BACKGROUND: Digital polymerase chain reaction (dPCR) is an accurate and sensitive molecular method that can be used in clinical diagnostic, prognostic, and predictive tests. The key component of the dPCR method is the partitioning of a single reaction into many thousands of droplets, nanochannels or other nano- or picoliter-sized reactions. This results in high enough sensitivity to detect rare nucleic acid targets and provides an absolute quantification of target sequences or alleles compared to other PCR-based methods...

An alarming rise in antimicrobial resistance worldwide has spurred efforts into the search for alternatives to antibiotic treatments. The use of bacteriophages, bacterial viruses harmless to humans, represents a promising approach with potential to treat bacterial infections (phage therapy). Recent advances in microscopy-based single-cell techniques have allowed researchers to develop new quantitative methodologies for assessing the interactions between bacteria and phages, especially the ability of phages to eradicate bacterial pathogen populations and to modulate growth of both commensal and pathogen populations...

BACKGROUND: The identification and quantification of viable Escherichia coli (E. coli) are important in multiple fields including the development of antimicrobial materials, water quality, food safety and infections diagnosis. However, the standard culture-based methods of viable E. coli detection suffer from long detection times (24 h) and complex operation, leaving the unmet requirement for fast assessing the efficiency of antimicrobial materials, early alerting the contamination of water and food, and immediately treatment of infections...

Clonal growth and competition underlie processes of key relevance in etiology, progression and therapy response across all cancers. Here, we demonstrate a novel experimental approach, based on multi-color, fluorescent tagging of cell nuclei, in combination with picoliter droplet deposition, to study the clonal dynamics in two- and three-dimensional cell cultures. The method allows for the simultaneous visualization and analysis of multiple clones in individual multi-clonal colonies, providing a powerful tool for studying clonal dynamics and identifying clonal populations with distinct characteristics...

Bacteria with antagonistic activity inhibit the growth of other bacteria through different mechanisms, including the production of antibiotics. As a result, these microorganisms are a prolific source of such compounds. However, searching for antibiotic-producing strains requires high-throughput techniques due to the vast diversity of microorganisms. Here, we screened and isolated bacteria with antagonistic activity against Escherichia coli expressing the green fluorescent protein ( E. coli -GFP). We used microfluidics to co-encapsulate and co-culture single cells from different strains within picoliter gel beads and analyzed them using fluorescence-activated cell sorting (FACS)...

The capacity to precisely pipette femtoliter volumes of liquid enables many applications, for example, to functionalize a nanoscale surface and manipulate fluids inside a single-cell. A pressure-controlled pipetting method is the most preferred, since it enables the widest range of working liquids. However, precisely controlling femtoliter volumes by pressure is challenging. In this work, a new concept is proposed that makes use of axisymmetrical phaseguides inside a microfluidic channel to pipette liquid in discrete steps of known volume...

Multiplexed and label-free mass spectrometry-based approaches with single-cell resolution have attributed surprising heterogeneity to presumed homogenous cell populations. Even though specialized experimental designs and instrumentation have demonstrated remarkable advances, the efficient sample preparation of single cells still lags. Here, we introduce the proteoCHIP, a universal option for single-cell proteomics sample preparation including multiplexed labeling up to 16-plex with high sensitivity and throughput...

Electrospray ionization (ESI) is one of the main techniques used in mass spectrometry (MS) of nonvolatile compounds. ESI is a disordered process, in which a large number of polydisperse droplets are projected from a fluctuating Taylor cone and jet protruding ESI emitter. Here, we disclose a system for sectioning electrospray plumes to discrete packets with millisecond and submillisecond lifetime, which are introduced to the MS orifice, one at a time. A high-speed camera was triggered at 10,000 frames per second to capture consecutive images of the electrospray packets transmitted to the mass spectrometer...

Droplets of alcohol-based formulations are common in applications from sanitizing sprays to printing inks. However, our understanding of the drying dynamics of these droplets on surfaces and the influence of ambient humidity is still very limited. Here, we report the drying dynamics of picoliter droplets of isopropyl alcohol deposited on a surface under controlled humidity. Condensation of water vapor in the ambient environment onto alcohol droplets leads to unexpectedly complex drying behavior. As relative humidity (RH) increases, we observed a variety of phenomena including enhanced spreading, nonmonotonic changes in the drying time, the formation of pancake-like shapes that suppress the coffee-ring effect, and the formation of water-rich films around an alcohol-rich drop...

The 3D colloidal assemblies composed of nanoparticles (NPs) are closely associated with optical properties such as photonic crystals, localized surface plasmon resonance, and surface-enhanced Raman scattering. However, research on their fabrication remains insufficient. Here, the femtoliter volume of a 3D colloidal assembly is shown, using the evaporation of a fine fountain pen. A nano-fountain pen (NPF) with a micrometer-level tip inner diameter was adopted for the fine evaporation control of the ink solvent...

An important approach to molecular diagnostics is integrating organized substances that provide complex molecular level responses to introduced chemical and biological agents with conditions that optimize and distinguish such responses. In this respect, liquid crystal dispersions are attractive components of molecular diagnostic tools. This paper analyzes a colloid system, containing a nematic liquid crystal as a dispersed phase, and aqueous surfactant and polymer solutions as the continuous phases. We applied a microfluidic approach for tuning orientation of liquid crystal molecules in picoliter droplets immobilized on microchannel walls...

The carbohydrate antigen 19-9 (CA 19-9) is the most commonly used biomarker in the clinical diagnosis of pancreatic cancer. Multiphoton nonlinear laser wave-mixing spectroscopy is presented as an ultrasensitive detection method for CA 19-9. Wave mixing is an optical absorption-based method, and hence, one can detect CA 19-9 without labels in their native form using compact ultraviolet (UV) lasers or labeled samples using a visible laser. The wave-mixing signal exhibits a quadratic dependence on the sample concentration, and hence, it is an ideal sensor to monitor small changes in the sample...

Capillary electrophoresis mass spectrometry (CE-MS) is an emerging analytical tool for microscale biological sample analysis that offers high separation resolution, low detection limit, and low sample consumption. We recently developed a novel microsampling device, "spray-capillary," for quantitative low-volume sample extraction (as low as 15 pL/s) and online CE-MS analysis. This platform can efficiently analyze picoliter samples (e.g., single cells) with minimal sample loss and no additional offline sample-handling steps...