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Keratins are an integral part of cell structure and function. Here, it is shown that ectopic expression of a truncated isoform of keratin 81 (tKRT81) in breast cancer is upregulated in metastatic lesions compared to primary tumors and patient‐derived circulating tumor cells, and is associated with more aggressive subtypes. tKRT81 physically interacts with keratin 18 (KRT18) and leads to changes in the cytosolic keratin intermediate filament network and desmosomal plaque formation. These structural changes are associated with a softer, more elastically deformable cancer cell with enhanced adhesion and clustering ability leading to greater in vivo lung metastatic burden. This work describes a novel biomechanical mechanism by which tKRT81 promotes metastasis, highlighting the importance of the biophysical characteristics of tumor cells. [ABSTRACT FROM AUTHOR]

Background: Understanding and countering the well-established negative health consequences of spaceflight remains a primary challenge preventing safe deep space exploration. Targeted/personalized therapeutics are at the forefront of space medicine strategies, and cross-species molecular signatures now define the 'typical' spaceflight response. However, a lack of direct genotype–phenotype associations currently limits the robustness and, therefore, the therapeutic utility of putative mechanisms underpinning pathological changes in flight. Methods: We employed the worm Caenorhabditis elegans as a validated model of space biology, combined with 'NemaFlex-S' microfluidic devices for assessing animal strength production as one of the most reproducible physiological responses to spaceflight. Wild-type and dys-1 (BZ33) strains (a Duchenne muscular dystrophy (DMD) model for comparing predisposed muscle weak animals) were cultured on the International Space Station in chemically defined media before loading second-generation gravid adults into NemaFlex-S devices to assess individual animal strength. These same cultures were then frozen on orbit before returning to Earth for next-generation sequencing transcriptomic analysis. Results: Neuromuscular strength was lower in flight versus ground controls (16.6% decline, p < 0.05), with dys-1 significantly more (23% less strength, p < 0.01) affected than wild types. The transcriptional gene ontology signatures characterizing both strains of weaker animals in flight strongly corroborate previous results across species, enriched for upregulated stress response pathways and downregulated mitochondrial and cytoskeletal processes. Functional gene cluster analysis extended this to implicate decreased neuronal function, including abnormal calcium handling and acetylcholine signaling, in space-induced strength declines under the predicted control of UNC-89 and DAF-19 transcription factors. Finally, gene modules specifically altered in dys-1 animals in flight again cluster to neuronal/neuromuscular pathways, suggesting strength loss in DMD comprises a strong neuronal component that predisposes these animals to exacerbated strength loss in space. Conclusions: Highly reproducible gene signatures are strongly associated with space-induced neuromuscular strength loss across species and neuronal changes in calcium/acetylcholine signaling require further study. These results promote targeted medical efforts towards and provide an in vivo model for safely sending animals and people into deep space in the near future. [ABSTRACT FROM AUTHOR]

Living longer without simultaneously extending years spent in good health ("health span") is an increasing societal burden, demanding new therapeutic strategies. Hydrogen sulfide (H2S) can correct disease-related mitochondrial metabolic deficiencies, and supraphysiological H2S concentrations can pro health span. However, the efficacy and mechanisms of mitochondrion-targeted sulfide delivery molecules (mtH2S) administered across the adult life course are unknown. Using a Caenorhabditis elegans aging model, we compared untargeted H2S (NaGYY4137, 100 µM and 100 nM) and mtH2S (AP39, 100 nM) donor effects on life span, neuromuscular health span, and mitochondrial integrity. H2S donors were administered from birth or in young/middle-aged animals (day 0, 2, or 4 postadulthood). RNAi pharmacogenetic interventions and transcriptomics/network analysis explored molecular events governing mtH2S donor-mediated health span. Developmentally administered mtH2S (100 nM) improved life/health span vs. equivalent untargeted H2S doses. mtH2S preserved aging mitochondrial structure, content (citrate synthase activity) and neuromuscular strength. Knockdown of H2S metabolism enzymes and FoxO/daf-16 prevented the positive health span effects of mtH2S, whereas DCAF11/wdr-23 - Nrf2/skn-1 oxidative stress protection pathways were dispensable. Health span, but not life span, increased with all adult-onset mtH2S treatments. Adult mtH2S treatment also rejuvenated aging transcriptomes by minimizing expression declines of mitochondria and cytoskeletal components, and peroxisome metabolism hub components, under mechanistic control by the elt-6/elt-3 transcription factor circuit. H2S health span extension likely acts at the mitochondrial level, the mechanisms of which dissociate from life span across adult vs. developmental treatment timings. The small mtH2S doses required for health span extension, combined with efficacy in adult animals, suggest mtH2S is a potential healthy aging therapeutic. [ABSTRACT FROM AUTHOR]

Neoadjuvant chemotherapy (NAC) can affect pathological complete response (pCR) in breast cancers; the resection that follows identifies patients with residual disease who are then offered second-line therapies. Circulating tumor cells (CTCs) and cancer-associated macrophage-like cells (CAMLs) in the blood can be used as potential biomarkers for predicting pCR before resection. CTCs are of epithelial origin that undergo epithelial-to-mesenchymal transition to become more motile and invasive, thereby leading to invasive mesenchymal cells that seed in distant organs, causing metastasis. Additionally, CAMLs in the blood of cancer patients are reported to either engulf or aid the transport of cancer cells to distant organs. To study these rare cancer-associated cells, we conducted a preliminary study where we collected blood from patients treated with NAC after obtaining their written and informed consent. Blood was collected before, during, and after NAC, and Labyrinth microfluidic technology was used to isolate CTCs and CAMLs. Demographic, tumor marker, and treatment response data were collected. Non-parametric tests were used to compare pCR and non-pCR groups. Univariate and multivariate models were used where CTCs and CAMLs were analyzed for predicting pCR. Sixty-three samples from 21 patients were analyzed. The median(IQR) pre-NAC total and mesenchymal CTC count/5 mL was lower in the pCR vs. non-pCR group [1(3.5) vs. 5(5.75); p = 0.096], [0 vs. 2.5(7.5); p = 0.084], respectively. The median(IQR) post-NAC CAML count/5 mL was higher in the pCR vs. non-pCR group [15(6) vs. 6(4.5); p = 0.004]. The pCR group was more likely to have >10 CAMLs post-NAC vs. non-pCR group [7(100%) vs. 3(21.4%); p = 0.001]. In a multivariate logistic regression model predicting pCR, CAML count was positively associated with the log-odds of pCR [OR = 1.49(1.01, 2.18); p = 0.041], while CTCs showed a negative trend [Odds Ratio (OR) = 0.44(0.18, 1.06); p = 0.068]. In conclusion, increased CAMLs in circulation after treatment combined with lowered CTCs was associated with pCR. [ABSTRACT FROM AUTHOR]

The model organism Caenorhabditis elegans is used in a variety of applications ranging from fundamental biological studies, to drug screening, to disease modeling, and to space-biology investigations. These applications rely on conducting whole-organism phenotypic assays involving animal behavior and locomotion. In this study, we report a 3D printed compact imaging platform (CIP) that is integrated with a smart-device camera for the whole-organism phenotyping of C. elegans. The CIP has no external optical elements and does not require mechanical focusing, simplifying the optical configuration. The small footprint of the system powered with a standard USB provides capabilities ranging from plug-and-play, to parallel operation, and to housing it in incubators for temperature control. We demonstrate on Earth the compatibility of the CIP with different C. elegans substrates, including agar plates, liquid droplets on glass slides and microfluidic chips. We validate the system with behavioral and thrashing assays and show that the phenotypic readouts are in good agreement with the literature data. We conduct a pilot study with mutants and show that the phenotypic data collected from the CIP distinguishes these mutants. Finally, we discuss how the simplicity and versatility offered by CIP makes it amenable to future C. elegans investigations on the International Space Station, where science experiments are constrained by system size, payload weight and crew time. Overall, the compactness, portability and ease-of-use makes the CIP desirable for research and educational outreach applications on Earth and in space. [ABSTRACT FROM AUTHOR]

Label-free technologies for isolating rare circulating cells in breast cancer patients are widely available; however, they are mostly validated on metastatic patient blood samples. Given the need to use blood-based biomarkers to inform on disease progression and treatment decisions, it is important to validate these technologies in non-metastatic patient blood samples. In this study, we specifically focus on a recently established label-free microfluidic technology Labyrinth and assess its capabilities to phenotype a variety of rare circulating tumor cells indicative of epithelial-to-mesenchymal transition as well as cancer-associated macrophage-like (CAML) cells. We specifically chose a patient cohort that is non-metastatic and selected to undergo neoadjuvant chemotherapy to assess the performance of the Labyrinth technology. We enrolled 21 treatment naïve non-metastatic breast cancer patients of various disease stages. Our results indicate that (i) Labyrinth microfluidic technology is successfully able to isolate different phenotypes of CTCs despite the counts being low. (ii) Invasive phenotypes of CTCs such as transitioning CTCs and mesenchymal CTCs were found to be present in high numbers in stage III patients as compared to stage II patients. (iii) As the total load of CTCs increased, the mesenchymal CTCs were found to be increasing. (iv) Labyrinth was able to isolate CAMLs with the counts being higher in stage III patients as compared to stage II patients. Our study demonstrates the ability of the Labyrinth microfluidic technology to isolate rare cancer-associated cells from the blood of treatment naïve non-metastatic breast cancer patients, laying the foundation for tracking oncogenic spread and immune response in patients undergoing neoadjuvant chemotherapy. [ABSTRACT FROM AUTHOR]

Encapsulation of cells inside microfluidic droplets is central to several applications involving cellular analysis. Although, theoretically the encapsulation statistics are expected to follow a Poisson distribution, experimentally this may not be achieved due to lack of full control of the experimental variables and conditions. Therefore, there is a need for automatic detection of droplets and cell count enumeration within droplets so a process control feedback to adjust experimental conditions can be implemented. In this study, we use a deep learning object detector called You Only Look Once (YOLO), an influential class of object detectors with several benefits over traditional methods. This paper investigates the application of both YOLOv3 and YOLOv5 object detectors in the development of an automated droplet and cell detector. Experimental data was obtained from a microfluidic flow focusing device with a dispersed phase of cancer cells. The microfluidic device contained an expansion chamber downstream of the droplet generator, allowing for visualization and recording of cell-encapsulated droplet images. In the procedure, a droplet bounding box is predicted, then cropped from the original image for the individual cells to be detected through a separate model for further examination. The system includes a production set for additional performance analysis with Poisson statistics while providing an experimental workflow with both droplet and cell models. The training set is collected and preprocessed before labeling and applying image augmentations, allowing for a generalizable object detector. Precision and recall were utilized as a validation and test set metric, resulting in a high mean average precision (mAP) metric for an accurate droplet detector. To examine model limitations, the predictions were compared to ground truth labels, illustrating that the YOLO predictions closely matched with the droplet and cell labels. Furthermore, it is demonstrated that droplet enumeration from the YOLOv5 model is consistent with hand counted ratios and the Poisson distribution, confirming that the platform can be used in real-time experiments for cell encapsulation optimization. [ABSTRACT FROM AUTHOR]

Inertial, size-based focusing was investigated in the microfluidic labyrinth device consisting of several U-shaped turns along with circular loops. Turns are associated with tight curvature and, therefore, induce strong Dean forces for separating particles; however, systematic studies exploring this possibility do not exist. We characterized the focusing dynamics of different-sized rigid particles, cancer cells, and white blood cells over a range of fluid Reynolds numbers R e f. Streak widths of the focused particle streams at all the turns showed intermittent fluctuations that were substantial for smaller particles and at higher R e f. In contrast, cell streaks were less prone to fluctuations. Computational fluid dynamics simulations revealed the existence of strong turn-induced Dean vortices, which help explain the intermittent fluctuations seen in particle focusing. Next, we developed a measure of pairwise separability to evaluate the quality of separation between focused streams of two different particle sizes. Using this, we assessed the impact of a single sharp turn on separation. In general, the separability was found to vary significantly as particles traversed the tight-curvature U-turn. Comparing the separability at the entry and exit sections, we found that turns either improved or reduced separation between different-sized particles depending on R e f. Finally, we evaluated the separability at the downstream expansion section to quantify the performance of the labyrinth device in terms of achieving size-based enrichment of particles and cells. Overall, our results show that turns are better for cell focusing and separation given that they are more immune to curvature-driven fluctuations in comparison to rigid particles. [ABSTRACT FROM AUTHOR]

A technique is described for continuous production of microparticles of variable size with new forms of anisotropy including alternating bond angles and configurable patchiness. The studies have shown that the particle's sequence is dynamically controlled through fluidic processing.

We report an experimental investigation on the effect of emulsifiers on the dripping-to-jetting transition and droplet size scaling in a co-flow microcapillary device. The emulsifiers studied include an oil-soluble surfactant Span 80, a water-soluble nanosheet, and their combination. We characterize the dispersed phase behaviors in the state space comprising the Weber number of the inner phase Wein and the capillary number of the outer phase Caout. Results show that the dripping-to-jetting (D-J) transition occurs at Wein ≈ O(1) for the no-emulsifier case and Wein ≈ 65 for the Span 80 system. In the case of the nanosheet, the D-J transition is dependent on concentration with Wein ≈ 0.08–5.6. For the combination system, depending on the nanosheet concentration, the critical Wein varies from 0.3 to 21. We explain that these emulsifier-dependent critical Wein result in terms of additional interfacial stresses arising due to surfactant transport and adsorbed nanosheets. Combining droplet size data from this study and literature, we find that in the dripping regime, the droplet diameter decreases with increasing Caout, yielding a scaling exponent of ≈−0.33 and a pre-factor that could be dependent on the type of emulsifier. In the jetting regime, the droplet diameter increases with the flow rate ratio of the two phases, with a scaling exponent of ≈ 0.37 that is independent of the type of emulsifier. The results from this work help to better understand the influence of surfactants and particulate emulsifiers in microfluidic emulsification. [ABSTRACT FROM AUTHOR]

Transcriptome‐based drug screening is emerging as a powerful tool to identify geroprotective compounds to intervene in age‐related disease. We hypothesized that, by mimicking the transcriptional signature of the highly conserved longevity intervention of FOXO3 (daf‐16 in worms) overexpression, we could identify and repurpose compounds with similar downstream effects to increase longevity. Our in silico screen, utilizing the LINCS transcriptome database of genetic and compound interventions, identified several FDA‐approved compounds that activate FOXO downstream targets in mammalian cells. These included the neuromuscular blocker atracurium, which also robustly extends both lifespan and healthspan in Caenorhabditis elegans. This longevity is dependent on both daf‐16 signaling and inhibition of the neuromuscular acetylcholine receptor subunit unc‐38. We found unc‐38 RNAi to improve healthspan, lifespan, and stimulate DAF‐16 nuclear localization, similar to atracurium treatment. Finally, using RNA‐seq transcriptomics, we identify atracurium activation of DAF‐16 downstream effectors. Together, these data demonstrate the capacity to mimic genetic lifespan interventions with drugs, and in doing so, reveal that the neuromuscular acetylcholine receptor regulates the highly conserved FOXO/DAF‐16 longevity pathway. [ABSTRACT FROM AUTHOR]

In this study, we report microfluidic shear rheology and wall-slip using the 3D-resolved flow kinematics obtained from digital holography microscopy (DHM). We computationally reconstruct the recorded holograms to visualize the tracer imbued flow volume in linear microchannels, followed by the implementation of particle tracking velocimetry (PTV) to quantitate spatially resolved velocity fields in 3D. In order to select optimal parameters for DHM-PTV characterization of viscoelastic fluids, we studied the effect of the hologram recording distance, seeding density, and particle size. Using the optimal parameters, we show quantitative characterization of the shear rheology from the velocity fields without any a priori assumptions of wall boundary conditions or constitutive equation. The viscosity vs shear rate data for Newtonian and polyethylene oxide (PEO) solutions could be measured in the range of ≈0.05 to 20 000 s−1 with just three input pressures using sample volumes as low as 20 µl. These data from holographic shear rheometry were found to be in good agreement with computational fluid dynamics simulations and macrorheometry. With respect to the wall-slip, we find that highly viscoelastic PEO solutions can show slip lengths in the order of few microns. Finally, we discuss holographic visualization of particle migration in microfluidic flows, which can limit flow field access, whereas at the same time provide a fingerprint of the suspending fluid rheology. [ABSTRACT FROM AUTHOR]

Primary cancers disseminate both single circulating tumor cells (CTCs) and CTC "clusters," the latter of which have been shown to demonstrate greater metastatic propensity and adverse impact on prognosis. Many devices developed to isolate single CTCs also capture CTC clusters, but there is translational potential for a platform specifically designed to isolate CTC clusters. Herein, we introduce our microfluidic device for isolating CTC clusters ("Microfluidic Isolation of CTC Clusters" or MICC), which is equipped with ∼10 000 trap chambers that isolate tumor cell clusters based on their large sizes and dynamic force balance against a pillar obstacle in the trap chamber. Whole blood is injected, followed by a wash step to remove blood cells and a final backflush to release intact clusters for downstream analysis. Using clusters from tumor cell-line and confocal microscopy, we verified the ability of the MICC platform to specifically capture tumor cell clusters in the trap chambers. Our flow rate optimization experiments identified 25 μl/min for blood injection, 100 μl/min as wash flow rate, and 300 μl/min as the release flow rate – indicating that 1 ml of whole blood can be processed in less than an hour. Under these optimal flow conditions, we assessed the MICC platform's capture and release performance using blood samples spiked with different concentrations of clusters, revealing a capture efficiency of 66%–87% and release efficiency of 76%–90%. The results from our study suggest that the MICC platform has the potential to isolate CTC clusters from cancer patient blood, enabling it for clinical applications in cancer management. [ABSTRACT FROM AUTHOR]

The relative velocity and extra pressure drop of a single vesicle flowing through a square microchannel are quantified via boundary element simulations, lubrication theory and microfluidic experiments. The vesicle is modelled as a fluid sac enclosed by an inextensible, fluidic membrane with a negligible bending stiffness. All results are parametrized in terms of the vesicle sphericity (i.e. the reduced volume) and flow confinement (i.e. the ratio of the vesicle radius to the channel hydraulic radius). Direct comparison is made to previous studies of vesicle flow through circular tubes, revealing several distinct features of the square-channel geometry. Firstly, fluid in the suspending medium bypasses the vesicle through the corners of the channel, which in turn reduces the dissipation created by the vesicle. Secondly, the absence of rotational symmetry about the channel axis permits surface circulation in the membrane (tank treading), which in turn reduces the vesicle's speed. At very high confinement, both theory and experiment indicate that the vesicle's speed can be reduced below the mean speed of the suspending fluid through this mechanism. Finally, the contact area for lubrication is greatly reduced in the square-duct geometry, which in turn weakens the stress singularity predicted by lubrication theory. This fact directly leads to a breakdown of the lubrication approximation at low flow confinement, as verified by comparison to boundary element simulations. Since the only distinct property assumed of the membrane is its ability to preserve surface area locally, it is expected that the results of this study are applicable to other types of soft particles with immobilized surfaces (e.g. Pickering droplets, gel beads and biological cells). [ABSTRACT FROM AUTHOR]

Long term spaceflight is associated with the loss of skeletal muscle mass and function. The Molecular Muscle Experiment (MME) seeks to identify the causes of muscle decline in space and test potential therapies to attenuate this in the microscopic worm, Caenorhabditis elegans. This is the first UK-led experiment in the almost two-decade history of the International Space Station. We therefore intend to complete significant and widespread educational outreach activities to promote interest in science, technology, engineering, and math (STEM), and to increase engagement with our space life science experiment. This paper describes three education outreach activities relating to MME that are suitable for use in the classroom: (i) observing normal and mutant worms; (ii) observing the effect of unloading (simulation of microgravity); and (iii) handling spaceflight hardware. Activity packs are provided at a starter and advanced level to support these activities. This paper also provides three posters that may be used as learning resources for educators. These posters provide information on: (i) why worms are used for research; (ii) spaceflight human physiology; and (iii) the specifics of the MME. Details of further planned engagement activities are outlined to increase the awareness of the MME. [ABSTRACT FROM AUTHOR]

Muscle strength is a functional measure of quality of life in humans. Declines in muscle strength are manifested in diseases as well as during inactivity, aging, and space travel. With conserved muscle biology, the simple genetic model C. elegans is a high throughput platform in which to identify molecular mechanisms causing muscle strength loss and to develop interventions based on diet, exercise, and drugs. In the clinic, standardized strength measures are essential to quantitate changes in patients; however, analogous standards have not been recapitulated in the C. elegans model since force generation fluctuates based on animal behavior and locomotion. Here, we report a microfluidics-based system for strength measurement that we call ‘NemaFlex’, based on pillar deflection as the nematode crawls through a forest of pillars. We have optimized the micropillar forest design and identified robust measurement conditions that yield a measure of strength that is independent of behavior and gait. Validation studies using a muscle contracting agent and mutants confirm that NemaFlex can reliably score muscular strength in C. elegans. Additionally, we report a scaling factor to account for animal size that is consistent with a biomechanics model and enables comparative strength studies of mutants. Taken together, our findings anchor NemaFlex for applications in genetic and drug screens, for defining molecular and cellular circuits of neuromuscular function, and for dissection of degenerative processes in disuse, aging, and disease. [ABSTRACT FROM AUTHOR]

Current lithography methods allow facile fabrication of microfluidic conduits where not only the shape of the bounding walls can be arbitrarily varied but also the internal conduit space can be laden with a variety of microstructures and wetting properties. This virtually infinite design space of microfluidic geometries brings in the challenge of how to quantify fluid resistance in a large number of microfluidic conduits, while maintaining operational simplicity. We report a versatile experimental technique referred to as microfluidic bypass manometry for measurement of pressure drop versus flow rate (ΔP–Q) relations in a parallelized manner. The technique involves introducing co-flowing laminar streams into a microfluidic network that contains a series of loops, where each loop is comprised of a test geometry and a bypass channel as a flow-rate sensing element. We optimize the network geometry and present operational considerations for microfluidic bypass manometry. To demonstrate the power of our technique, we used single-phase fluids and measured ΔP–Q relations simultaneously for forty test geometries ranging from linear to contraction–expansion to serpentine to pillar-laden microchannels. To expand the capabilities of the method, we measured ΔP–Q relations for similar-sized oil droplets trapped in microcavities where the cavity geometry spans from prisms of 3–10 sides to circular disks. We found in all cases, the ΔP–Q relation is nonlinear and the flow resistance of droplets is sensitive to confinement. At high flow rates, the drop resistance depends on the cavity geometry and is higher in a triangular prism compared to a circular disk. We compared the measured flow resistance of single-phase fluids and droplets in different microfluidic geometries to that from computational fluid dynamics simulations and found them to be in excellent agreement. Given the simplicity and versatility of the microfluidic bypass manometry method, we anticipate that it may find broad application in several areas including design of lab-on-chip devices, laminar drag reduction and mechanics of deformable particles. [ABSTRACT FROM AUTHOR]

Cancer metastasis involves the dissemination of tumor cells into distant organ sites, with the circulatory system playing an important role in the dissemination process. SP sp [1] SP sp Circulating tumor cells (CTCs), circulating tumor DNA (ctDNA), and exosomes have been found in the blood of cancer patients, making these circulating biomarkers a rich source of diagnostic and prognostic information for personalized cancer management. SP sp [2] SP sp The development and advancement of microfluidic approaches to isolate and characterize these circulating biomarkers offers tremendous potential for translating these liquid biopsy approaches into the clinic. 10.1158/0008-5472.CAN-17-2748 9 V. Murlidharet al., "Poor prognosis indicated by venous circulating tumor cell clusters in early-stage lung cancers", Cancer Res. 77(18), 5194-5206 (2017). The article by Kamyabi I et al. i SP sp [10] SP sp reports an innovative microfluidic system that can process 1 ml of undiluted whole blood within 1 h for isolation and recovery of tumor cell clusters. [Extracted from the article]

We investigate the flow resistance of a droplet trapped at a constriction in a microcavity located at a microchannel bifurcation as a function of system parameters including capillary number, drop confinement, and viscosity ratio. Using a combination of experiments and volume-of-fluid numerical simulations, we measure the hydrodynamic resistance of the trapped drop and connect it to drop deformation in the microcavity. For drop sizes smaller than the microcavity, we observe a bistable behavior in terms of the resistance of the trapped drop as a function of capillary number. For these underfilled drops, we find that the resistance is low at small capillary number (Ca < 10) and jumps to high resistance at a threshold capillary number. For drops equal to the microcavity size, we observe that the bistability vanishes and the drop resistance is of similar magnitude as that of underfilled drops at large capillary number. To explain these findings, we use confocal microscopy and simulations to obtain three-dimensional views of the drop deformation and continuous phase fluid in the microcavity. We observe that the low resistance is due to negligible drop deformation and unobstructed flow of continuous phase through the constriction. The high resistance is due to the drop interface protruding into the constriction restricting the flow of continuous phase through the gutters. Taken together, our results indicate that a trapped drop at a bifurcation can act as a nonlinear resistor and could be potentially used as a soft switch to control droplet trajectories in microfluidic devices. [ABSTRACT FROM AUTHOR]

We introduce inline digital holographic microscopy (in-line DHM) as a label-free technique for detecting tumor cells in blood. The optimized DHM platform fingerprints every cell flowing through a microchannel at 10 000 cells per second, based on three features – size, maximum intensity and mean intensity. To identify tumor cells in a background of blood cells, we developed robust gating criteria using machine-learning approaches. We established classifiers from the features extracted from 100 000-cell training sets consisting of red blood cells, peripheral blood mononuclear cells and tumor cell lines. The optimized classifier was then applied to targeted features of a single cell in a mixed cell population to make quantitative cell-type predictions. We tested our classification system with tumor cells spiked at different levels into a background of lysed blood that contained predominantly peripheral blood mononuclear cells. Results show that our holographic screening method can readily detect as few as 10 tumor cells per mL, and can identify tumor cells at a false positive rate of at most 0.001%. This purely optical approach obviates the need for antibody labeling and allows large volumes of sample to be quickly processed. Moreover, our in-line DHM approach can be combined with existing circulation tumor cell enrichment strategies, making it a promising tool for label-free analysis of liquid-biopsy samples. [ABSTRACT FROM AUTHOR]

We used volume-of-fluid (VOF) method to perform three-dimensional numerical simulations of droplet formation of Newtonian fluids in microfluidic T-junction devices. To evaluate the performance of the VOF method we examined the regimes of drop formation and determined droplet size as a function of system parameters. Comparison of the simulation results with four sets of experimental data from the literature showed good agreement, validating the VOF method. Motivated by the lack of adequate studies investigating the influence of viscosity ratio (λ) on the generated droplet size, we mapped the dependence of drop volume on capillary number (0.001 < Ca < 0.5) and viscosity ratio (0.01 < λ < 15). We find that for all viscosity ratios investigated, the droplet size decreases with increase in the capillary number. However, the reduction in the droplet size with the capillary number is stronger for λ < 1 than for λ > 1. In addition, we find that at a given capillary number, the size of droplets does not vary appreciably when λ < 1, while it increases when λ > 1. We develop an analytical model for predicting the droplet size that includes a viscosity-dependent breakup time for the dispersed phase. This improved model successfully predicts the effects of the viscosity ratio observed in simulations. Results from this study are useful for the design of lab-on-chip technologies and manufacture of microfluidic emulsions, where there is a need to know how system parameters influence the droplet size. [ABSTRACT FROM AUTHOR]