Microfluidic Chamber Research Articles

Observation of Enantiomeric Switching of Individual Plasmonic Metamolecules.

Active plasmonic metamolecules under microscopic observation are promising for optical reporters in single molecule sensing applications. While self-assembled reconfigurable chiral plasmonic metamolecules can be conveniently engineered with sensing functionalities, their observation is usually based on ensemble measurements, where the chiroptical response of enantiomers tend to cancel each other in ensemble circular dichroism. Herein, we demonstrate microscopic observation of enantiomeric switching of individual active DNA origami-assembled plasmonic metamolecules. The metamolecules are immobilized on a glass substrate in a microfluidic chamber, in which the plasmonic metamolecule can maintain their activities upon certain local stimuli as in solution. In circular differential scattering, two enantiomeric states controlled by the strand-displacement reaction display opposite spectral signals to each other, representing successful enantiomeric switching of the chirality. Moreover, in a close-to-racemic mixture of chiral metamolecules controlled by pH-sensitive strands, the coexistence of enantiomeric individuals, which is concealed in ensemble measurements, is clearly identified.

Enhanced and stabilized mesenchymal stem cell growth inside plasma pre-treated and collagen-coated PDMS-based microfluidic chambers

Poly(dimethylsiloxane) (PDMS) – based microfluidic devices, let them be lab-on-a-chip or organ-on-a-chip platforms, have been extensively utilized in biomedical research, due to the versatility, rapid prototyping, optical transparency and ease of fabrication. Despite its numerous advantages, PDMS is inherently hydrophobic, which compromises cell adhesion and proliferation, given the direct correlation between surface properties and cell viability. Various strategies have been followed for the functionalization of PDMS surface, from which extracellular matrix (ECM) protein immobilization appears to be more compatible with cell-related applications. In this context, current studies propose multi-step, tedious and occasionally environmentally hazardous procedures of PDMS chemical functionalization that are additionally not easily employed in microfluidic systems. To overcome these limitations, this work suggests air plasma treatment as the single-step PDMS activation procedure for the immobilization of type I collagen, and is one of the few to actually be applied inside microfluidic devices, at the integral fabrication step of sealing, to study cell sustainability. Stable hydrophilization of the plasma-treated and collagen-coated PDMS surfaces was verified for a time period of 7 days in dry state, achieving the highest reported wettability. Moreover, mesenchymal stem cells (MSCs) were cultured in as-treated 3D microchambers for 5 days, when they reached complete confluency, one of the highest reported in literature. The results revealed the potential of plasma treatment followed shortly after by collagen layering to support stabilized and long-term cell culture in PDMS-based microfluidic devices. The herein proposed PDMS surface functionalization method can be used to create a stable PDMS-collagen composite layer in organ-on-a-chip platforms.

Dynamic control of high-voltage actuator arrays by light-pattern projection on photoconductive switches

The ability to control high-voltage actuator arrays relies, to date, on expensive microelectronic processes or on individual wiring of each actuator to a single off-chip high-voltage switch. Here we present an alternative approach that uses on-chip photoconductive switches together with a light projection system to individually address high-voltage actuators. Each actuator is connected to one or more switches that are nominally OFF unless turned ON using direct light illumination. We selected hydrogenated amorphous silicon (a-Si:H) as our photoconductive material, and we provide a complete characterization of its light to dark conductance, breakdown field, and spectral response. The resulting switches are very robust, and we provide full details of their fabrication processes. We demonstrate that the switches can be integrated into different architectures to support both AC and DC-driven actuators and provide engineering guidelines for their functional design. To demonstrate the versatility of our approach, we demonstrate the use of the photoconductive switches in two distinctly different applications—control of µm-sized gate electrodes for patterning flow fields in a microfluidic chamber and control of cm-sized electrostatic actuators for creating mechanical deformations for haptic displays.

Parallel Dielectrophoretic Capture, Isolation, and Electrical Lysis of Individual Breast Cancer Cells to Assess Variability in Enzymatic Activity.

Tumor cell heterogeneity drives disease progression and response to therapy, and therefore, there is a need for single-cell analysis methods. In this paper, we present an integrated, scalable method to analyze enzymatic activity in many individual cancer cells at once. The reported method uses dielectrophoresis (DEP) to selectively capture tumor cells at wireless electrodes aligned to an overlying array of cell-sized micropockets. Following hydrodynamic transfer of the captured cells into microfluidic chambers, the chambers are fluidically isolated and sealed with a hydrophobic ionic liquid, which possesses sufficient conductivity to allow for subsequent electrical lysis of the cells to access their contents for enzymatic assay. The wireless electrodes have an interlocking spiral design that ensures successful electrical lysis regardless of the location of the cell within the chamber. Here, breast cancer cells are assessed for β-galactosidase through its activation of a fluorogenic substrate. A key point is that the fluorogenic assay solution was optimized to allow for dielectrophoretic cell capture, thereby obviating the need for a solution exchange step. Our approach has several distinct advantages including a high rate of single-cell capture, a capture efficiency that is independent of the dimensions of the reaction chambers, no need for mechanical closure of reaction volumes, and no observed cross-talk. In this study, first, the steps of cell capture, transfer, and lysis are established on this platform in the presence of the optimized assay solution. We then quantify the increase in fluorescence intensity obtained over the duration of the enzymatic assay of individual cells. Finally, this method is applied to the analysis of β-galactosidase activity in 258 individual MDA-MB-231 breast cancer cells, revealing heterogeneity in expression of this enzyme in this cell line. We expect that the adaptability of this method will allow for expanded studies of single-cell enzymatic expression and activity. This will in turn open avenues of research into cancer cell heterogeneity in metabolism, invasiveness, and drug response. The ability to study these features of cancer at the single-cell level raises the possibility for treatment plans tailored to target the specific combinations of cell subpopulations present in tumors. Furthermore, we expect that this method can be adapted to uses outside of cancer research, such as studies of neuron metabolism, pathogenesis in bacteria, and stem cell development.

Nuclei Isolation from Mouse Cardiac Progenitor Cells for Epigenome and Gene Expression Profiling at Single-Cell Resolution.

The developing heart is a complex structure containing various progenitor cells controlled by complex regulatory mechanisms. The examination of the gene expression and chromatin state of individual cells allows the identification of the cell type and state. Single-cell sequencing approaches have revealed a number of important characteristics of cardiac progenitor cell heterogeneity. However, these methods are generally restricted to fresh tissue, which limits studies with diverse experimental conditions, as the fresh tissue must be processed at once in the same run to reduce the technical variability. Therefore, easy and flexible procedures to produce data from methods such as single-nucleus RNA sequencing (snRNA-seq) and the single-nucleus assay for transposase-accessible chromatin with high-throughput sequencing (snATAC-seq) are needed in this area. Here, we present a protocol to rapidly isolate nuclei for subsequent single-nuclei dual-omics (combined snRNA-seq and snATAC-seq). This method allows the isolation of nuclei from frozen samples of cardiac progenitor cells and can be combined with platforms that use microfluidic chambers.

Development of cloth-based microfluidic devices for rapid determination of histamine in fish and fishery products.

A cloth-based analytical device combined with electrochemiluminescence detection (CAD-ECL) was described for rapid determination of histamine (HA). The CAD device was produced by screen-printing a conductive carbon ink onto a patterned hydrophobic electrochemical microfluidic chamber to fabricate the three-carbon electrode system on a single hydrophilic cloth. The introduction of carbon nanodots linked to chitosan on the working carbon electrode surface enhanced the catalytic performance and overcame the resistance of thecotton fibermaterial. On this basis, the enhancement of the electrochemiluminescence (ECL) signal ofthe tris(2,2'-bipyridyl) ruthenium(II) complex, caused by HA, was observed in a phosphate buffer solution at pH 7.6. The proposed CAD-ECL sensor was successfully applied tothe quantification of HA in fish and fishery samples with good linearity between ECL intensity and the logarithm of HA concentration in the range 1.0 to 1000.0µg L-1 with a low detection limit of 0.82µg L-1.

Optimization design of interdigitated microelectrodes with an insulation layer on the connection tracks to enhance efficiency of assessment of the cell viability

BackgroundMicroelectrical Impedance Spectroscopy (µEIS) is a tiny device that utilizes fluid as a working medium in combination with biological cells to extract various electrical parameters. Dielectric parameters of biological cells are essential parameters that can be extracted using µEIS. µEIS has many advantages, such as portability, disposable sensors, and high-precision results.ResultsThe paper compares different configurations of interdigitated microelectrodes with and without a passivation layer on the cell contact tracks. The influence of the number of electrodes on the enhancement of the extracted impedance for different types of cells was provided and discussed. Different types of cells are experimentally tested, such as viable and non-viable MCF7, along with different buffer solutions. This study confirms the importance of µEIS for in vivo and in vitro applications. An essential application of µEIS is to differentiate between the cells’ sizes based on the measured capacitance, which is indirectly related to the cells’ size. The extracted statistical values reveal the capability and sensitivity of the system to distinguish between two clusters of cells based on viability and size.ConclusionA completely portable and easy-to-use system, including different sensor configurations, was designed, fabricated, and experimentally tested. The system was used to extract the dielectric parameters of the Microbeads and MCF7 cells immersed in different buffer solutions. The high sensitivity of the readout circuit, which enables it to extract the difference between the viable and non-viable cells, was provided and discussed. The proposed system can extract and differentiate between different types of cells based on cells’ sizes; two other polystyrene microbeads with different sizes are tested. Contamination that may happen was avoided using a Microfluidic chamber. The study shows a good match between the experiment and simulation results. The study also shows the optimum number of interdigitated electrodes that can be used to extract the variation in the dielectric parameters of the cells without leakage current or parasitic capacitance.

Pressure-controlled microfluidics for automated single-molecule sample preparation

Sample preparation is a crucial step in single-molecule experiments and involves passivating the microfluidic sample chamber, immobilizing the molecules, and setting experimental buffer conditions. The efficiency of the experiment depends on the quality and speed of sample preparation, which is often performed manually and relies on the experience of the experimenter. This can result in inefficient use of single-molecule samples and time, especially for high-throughput applications. To address this, a pressure-controlled microfluidic system is proposed to automate single-molecule sample preparation. The hardware is based on microfluidic components from ElveFlow and is designed to be cost-effective and adaptable to various microscopy applications. The system includes a reservoir pressure adapter and a reservoir holder designed for additive manufacturing. Two flow chamber designs Ibidi µ-slide and Grace Bio-Labs HybriWell chamber are characterized, and the flow characteristics of the liquid at different volume flow rates V̇ are simulated using CFD-simulations and compared to experimental and theoretical values. The goal of this work is to establish a straightforward and robust system for single-molecule sample preparation that can increase the efficiency of experiments and reduce the bottleneck of manual sample preparation, particularly for high-throughput applications.

Botulinum neurotoxin A modulates the axonal release of pathological tau in hippocampal neurons

Pathological tau aggregates propagate across functionally connected neuronal networks in human neurodegenerative pathologies, such as Alzheimer's disease. However, the mechanism underlying this process is poorly understood. Several studies have showed that tau release is dependent on neuronal activity and that pathological tau is found in the extracellular space in free form, as well as in the lumen of extracellular vesicles. We recently showed that metabotropic glutamate receptor activity and SNAP25 integrity modulate the release of pathological tau from human and mouse synaptosomes. Here, we have leveraged botulinum neurotoxins (BoNTs), which impair neurotransmitter release by cleaving specific synaptic SNARE proteins, to dissect molecular mechanisms related to tau release at synapses. In particular, we have tested the effect of botulinum neurotoxin A (BoNT/A) on the synaptic release of tau in primary mouse neurons. Hippocampal neurons were grown in microfluidic chambers and transduced with lentiviruses expressing human tau (hTau). We found that neuronal stimulation significantly increases the release of mutant hTau, whereas wild-type hTau is unaffected. Importantly, BoNT/A blocks mutant hTau release, indicating that this process is controlled by SNAP25, a component of the SNARE complex, in intact neurons. These results suggest that BoNTs are potent tools to study the spreading of pathological proteins in neurodegenerative diseases and could play a central role in identifying novel molecular targets for the development of therapeutic interventions to treat tauopathies.

Zinc Oxide-Based Nanoformulation Zinkicide Mitigates the Xylem-Limited Pathogen Xylella fastidiosa in Tobacco and Southern Highbush Blueberry.

The xylem-limited pathogen Xylella fastidiosa causes severe economic losses worldwide, and no effective antimicrobial disease management options are available. The goal of this study was to evaluate the efficacy of a novel ZnO-based nanoparticle formulation, Zinkicide TMN110 (ZnK), against X. fastidiosa in vitro and in planta. In vitro, minimum bactericidal concentration (MBC) of ZnK analyzed in Pierce's Disease 2 medium was estimated at approximately 60 ppm. Time-kill kinetics assay showed a 100% reduction of culturable X. fastidiosa in less than 1 h after ZnK treatment. Microfluidic chambers assays showed that ZnK also inhibits X. fastidiosa cell aggregation and growth under flow conditions. Phytotoxicity assessments in the greenhouse demonstrated that ZnK can be applied as a soil drench in 50 ml at 500 ppm/plant/week up to four times to tobacco and blueberry without causing visible damage. ZnK was also evaluated for disease control in the greenhouse using tobacco infected with X. fastidiosa subsp. fastidiosa strain TemeculaL. ZnK soil drench weekly applications at concentrations of 500 followed by 1,000 ppm (500/1,000) and 500/500/1,000 ppm (in 50 ml each), reduced X. fastidiosa populations by >2 to 3 log10 units and disease severity by approximately 57 and 76%, respectively, compared with the untreated control. Similarly, when blueberry plants infected with X. fastidiosa subsp. multiplex strain AlmaEm3 were soil drenched with ZnK at concentrations 1,000/1,000 ppm and 1,000/1,000/500 ppm (in 200 ml each), the bacterial population was reduced by approximately 1 to 2 log10 units, and disease severity decreased by approximately 39 and 43%, respectively. Overall, this study shows antibacterial activity of ZnK against X. fastidiosa and its effectiveness in plants to reduce disease symptoms under controlled conditions.

Effects of the circulating environment of COVID-19 on platelet and neutrophil behavior.

Thromboinflammatory complications are well described sequalae of Coronavirus Disease 2019 (COVID-19), and there is evidence of both hyperreactive platelet and inflammatory neutrophil biology that contributes to the thromoinflammatory milieu. It has been demonstrated in other thromboinflammatory diseases that the circulating environment may affect cellular behavior, but what role this environment exerts on platelets and neutrophils in COVID-19 remains unknown. We tested the hypotheses that 1) plasma from COVID-19 patients can induce a prothrombotic platelet functional phenotype, and 2) contents released from platelets (platelet releasate) from COVID-19 patients can induce a proinflammatory neutrophil phenotype. We treated platelets with COVID-19 patient and disease control plasma, and measured their aggregation response to collagen and adhesion in a microfluidic parallel plate flow chamber coated with collagen and thromboplastin. We exposed healthy neutrophils to platelet releasate from COVID-19 patients and disease controls and measured neutrophil extracellular trap formation and performed RNA sequencing. We found that COVID-19 patient plasma promoted auto-aggregation, thereby reducing response to further stimulation ex-vivo. Neither disease condition increased the number of platelets adhered to a collagen and thromboplastin coated parallel plate flow chamber, but both markedly reduced platelet size. COVID-19 patient platelet releasate increased myeloperoxidasedeoxyribonucleic acid complexes and induced changes to neutrophil gene expression. Together these results suggest aspects of the soluble environment circulating platelets, and that the contents released from those neutrophil behavior independent of direct cellular contact.

Reliable cell retention of mammalian suspension cells in microfluidic cultivation chambers

Microfluidic cultivation, with its high level of environmental control and spatio-temporal resolution of cellular behavior, is a well-established tool in today’s microfluidics. Yet, reliable retention of (randomly) motile cells inside designated cultivation compartments still represents a limitation, which prohibits systematic single-cell growth studies. To overcome this obstacle, current approaches rely on complex multilayer chips or on-chip valves, which makes their application for a broad community of users infeasible. Here, we present an easy-to-implement cell retention concept to withhold cells inside microfluidic cultivation chambers. By introducing a blocking structure into a cultivation chamber’s entrance and nearly closing it, cells can be manually pushed into the chamber during loading procedures but are unable to leave it autonomously in subsequent long-term cultivation. CFD simulations as well as trace substance experiments confirm sufficient nutrient supply within the chamber. Through preventing recurring cell loss, growth data obtained from Chinese hamster ovary cultivation on colony level perfectly match data determined from single-cell data, which eventually allows reliable high throughput studies of single-cell growth. Due to its transferability to other chamber-based approaches, we strongly believe that our concept is also applicable for a broad range of cellular taxis studies or analyses of directed migration in basic or biomedical research.

On‐chip purification of tetracycline from food matrices

AbstractAntibiotics are widespread both to treat human and animal diseases and to improve growth in food animals. However, their overuse in food‐producing animals has led to critical issues for human health, such as the direct toxicity and the development of antibiotic‐resistant bacterial strains. Therefore, the identification of traces of antibiotic in food before entering the market has became extremely important and brought out the need for novel bioanalytical methods and protocols. To meet this need, here, a microfluidic system was set up for the purification of tetracycline (TC) from raw milk, honey, and eggs. The system is based on the use of magnetic beads exposing copper ions, which are loaded in a microfluidic chamber and actuated by a ready‐made device. Tetracycline is captured by copper ions present on the microbeads, purified from the unwanted raw material present in the initial sample and recovered by a suitable elution solution. Different elution solutions were tested and results were evaluated by X‐ray photoelectron spectroscopy (XPS) and visible spectrometry. The microfluidic system was successfully employed for the purification of TC from raw milk, honey, and eggs after an initial dilution in buffer. The overall protocol was, therefore, demonstrated to efficiently purify tetracycline, laying the bases for a future implementation of in‐field on‐chip tests.

Paclitaxel effects on axonal localization and vesicular trafficking of NaV1.8.

Patients treated with paclitaxel (PTX) or other antineoplastic agents can experience chemotherapy-induced peripheral neuropathy (CIPN), a debilitating side effect characterized by numbness and pain. PTX interferes with microtubule-based transport, which inhibits tumor growth via cell cycle arrest but can also affect other cellular functions including trafficking of ion channels critical to transduction of stimuli by sensory neurons of the dorsal root ganglia (DRG). We examined the effects of PTX on voltage-gated sodium channel NaV1.8, which is preferentially expressed in DRG neurons, using a microfluidic chamber culture system and chemigenetic labeling to observe anterograde channel transport to the endings of DRG axons in real time. PTX treatment increased the numbers of NaV1.8-containing vesicles traversing the axons. Vesicles in PTX-treated cells exhibited greater average velocity, along with shorter and less frequent pauses along their trajectories. These events were paralleled by greater surface accumulation of NaV1.8 channels at the distal ends of DRG axons. These results were consistent with observations that NaV1.8 is trafficked in the same vesicles containing NaV1.7 channels, which are also involved in pain syndromes in humans and are similarly affected by PTX treatment. However, unlike Nav1.7, we did not detect increased NaV1.8 current density measured at the neuronal soma, suggesting a differential effect of PTX on trafficking of NaV1.8 in soma versus axonal compartments. Therapeutic targeting of axonal vesicular traffic would affect both Nav1.7 and Nav1.8 channels and increase the possibilities of alleviating pain associated with CIPN.

Abstract TMP116: A Novel Microfluidic Model Of Ischemic Stroke Recapitulating Arterial Occlusion And Prolonged Retraction

To our knowledge, an ex vivo model of thrombotic occlusion that (1) mimics arterial setting, (2) allows true occlusion, and (3) drug delivery without disturbing the thrombus does not exist. A platform with these capabilities would allow for high-throughput mechanistic study of thrombosis and thrombolysis. Here we present a microfluidic system that achieves these goals, providing a biofidelic ex vivo platform to study ischemic stroke. A mold with an upstream Y and a region of stenosis was fabricated and used to manufacture microfluidic chambers. Channels were coated with collagen at the stenosis. Heparinized blood was collected from healthy human donors (N=3) and platelets were labelled with fluorescent anti-CD41. Samples were perfused at 10000/s and outflow mass collected on a scale. Occlusion was indicated by mass plateau, and thrombi were left to retract in situ for 0 or 6 hrs. Kinetic immunofluorescent images were captured and used to derive surface area and the image profile by averaging MFI in each pixel column. Sample from the same donor dosed with 0.7 or 7nM alteplase (ALT) was perfused using the Y inlet using a constant pressure head (1000/s; mimicking middle cerebral artery) for 120 min. After 6 hours of post-occlusion retraction, thrombi substantially remodeled. The MFI curves (Fig 1) demonstrate even distribution of MFI post-occlusion that shifts to a central peak after 6 hr. Dosed sample from the same donor was able to be delivered upstream of the thrombus without disruption. Immediate reperfusion resulted in a reduction in thrombus surface area of (mean (SD)) 2% (10%) with 0.7nM ALT, 23% (7%) with 7nM, and 15% (1%) with vehicle. Reperfusion after 6 hrs resulted in a reduction of 14% (14%) with 0.7nM, 45% (12%) with 7nM, and 17% (8%) with vehicle. We have established an ex vivo high throughput system capable of recapitulating arterial occlusion, thrombus retraction in situ , drug delivery, and thrombolysis. This system can be used for mechanistic inquiry and drug discovery.

Multifunctional acoustofluidic centrifuge device using tri-symmetrical design for particle enrichment and separation and multiphase microflow mixing

Effective enrichment and separation of particles and mixing of multiphase microflows can be technically challenging in microfluidic sample processing and analysis. Acoustofluidic techniques have advantages such as high biocompatibility, versatility, and simple device design to meet these demands. Here, we present a novel tri-symmetrical acoustofluidic centrifuge based on traveling surface acoustic waves (TSAWs), achieving particle enrichment and bidirectional size-based separation, pushing away the larger particles while enriching the smaller ones, in a rapid time (<6 s). A microfluidic chamber was designed for further extraction of treated particles and cells. Experimental results show that our device can achieve enrichment of particles in a range of sizes and high purity separation (e.g. >90 % in 5 µm and 40 µm sample), demonstrating its sample processing capabilities. The acoustic treatment did not affect cells (consistent with the control group above 80 %). TSAW-induced acoustic streaming enabled efficient mixing (mixing efficiency χ >85 %) of fluorescent multi-phase microflows within a short time (<0.5 s). The underlying physical mechanisms have been verified by numerical simulations.

Failure to Thrive: Impaired BDNF Transport along the Cortical–Striatal Axis in Mouse Q140 Neurons of Huntington’s Disease

Boosting trophic support to striatal neurons by increasing levels of brain-derived neurotrophic factor (BDNF) has been considered as a target for therapeutic intervention for several neurodegenerative diseases, including Huntington's disease (HD). To aid in the implementation of such a strategy, a thorough understanding of BDNF cortical-striatal transport is critical to help guide its strategic delivery. In this manuscript, we investigate the dynamic behavior of BDNF transport along the cortical-striatal axis in Q140 primary neurons, a mouse model for HD. We examine this by using single-molecule labeling of BDNF conjugated with quantum dots (QD-BDNF) to follow the transport along the cortical-striatal axis in a microfluidic chamber system specifically designed for the co-culture of cortical and striatal primary neurons. Using this approach, we observe a defect of QD-BDNF transport in Q140 neurons. Our study demonstrates that QD-BDNF transport along the cortical-striatal axis involves the impairment of anterograde transport within axons of cortical neurons, and of retrograde transport within dendrites of striatal neurons. One prominent feature we observe is the extended pause time of QD-BDNF retrograde transport within Q140 striatal dendrites. Taken together, these finding support the hypothesis that delinquent spatiotemporal trophic support of BDNF to striatal neurons, driven by impaired transport, may contribute to the pathogenesis of HD, providing us with insight into how a BDNF supplementation therapeutic strategy may best be applied for HD.

Convection Confounds Measurements of Osmophoresis for Lipid Vesicles in Solute Gradients.

Lipid vesicles immersed in solute gradients are predicted to migrate from regions of high to low solute concentration due to osmotic flows induced across their semipermeable membranes. This process─known as osmophoresis─is potentially relevant to biological processes such as vesicle trafficking and cell migration; however, there exist significant discrepancies (several orders of magnitude) between experimental observations and theoretical predictions for the vesicle speed. Here, we seek to reconcile predictions of osmophoresis with observations of vesicle motion in osmotic gradients. We prepare quasi-steady solute gradients in a microfluidic chamber using density-matched solutions of sucrose and glucose to eliminate buoyancy-driven flows. We quantify the motions of giant DLPC vesicles and Brownian tracer particles in such gradients using Bayesian analysis of particle tracking data. Despite efforts to mitigate convective flows, we observe directed motion of both lipid vesicles and tracer particles in a common direction at comparable speeds of order 10 nm/s. These observations are not inconsistent with models of osmophoresis, which predict slower motion at ca. 1 nm/s; however, experimental uncertainty and the confounding effects of fluid convection prohibit a quantitative comparison. In contrast to previous reports, we find no evidence for anomalously fast osmophoresis of lipid vesicles when fluid convection is mitigated and quantified. We discuss strategies for enhancing the speed of osmophoresis using high permeability membranes and geometric confinement.

Real-time continuous monitoring of dynamic concentration profiles studied with biosensing by particle motion.

Real-time monitoring-and-control of biological systems requires lab-on-a-chip sensors that are able to accurately measure concentration-time profiles with a well-defined time delay and accuracy using only small amounts of sampled fluid. Here, we study real-time continuous monitoring of dynamic concentration profiles in a microfluidic measurement chamber. Step functions and sinusoidal oscillations of concentrations were generated using two pumps and a herringbone mixer. Concentrations in the bulk of the measurement chamber were quantified using a solution with a dye and light absorbance measurements. Concentrations near the surface were measured using a reversible cortisol sensor based on particle motion. The experiments show how the total time delay of the real-time sensor has contributions from advection, diffusion, reaction kinetics at the surface and signal processing. The total time delay of the studied real-time cortisol sensor was ∼90 seconds for measuring 63% of the concentration change. Monitoring of sinusoidal cortisol concentration-time profiles showed that the sensor has a low-pass frequency response with a cutoff frequency of ∼4 mHz and a lag time of ∼60 seconds. The described experimental methodology paves the way for the development of monitoring-and-control in lab-on-a-chip systems and for further engineering of the analytical characteristics of real-time continuous biosensors.

Acoustofluidic large-scale mixing for enhanced microfluidic immunostaining for tissue diagnostics.

The usage of microfluidics for automated and fast immunoassays has gained a lot of interest in the last decades. This integration comes with certain challenges, like the reconciliation of laminar flow patterns of micro-scale systems with diffusion-limited mass transport. Several methods have been investigated to enhance microfluidic mixing in microsystems, including acoustic-based fluidic streaming. Here, we report both by numerical simulation and experiments on the beneficiary effect of acoustic agitation on the uniformity of immunostaining in large-size and thin microfluidic chambers. Moreover, we investigate by numerical simulation the impact of reducing the incubation times and the concentrations of the biochemical detection reagents on the obtained immunoassay signal. Finally, acoustofluidic mixing was successfully used to reduce by 80% the incubation time of the Her2 (human epidermal growth factor receptor 2) and CK (cytokeratins) biomarkers for the spatial immunostaining of breast cancer cell pellets, or reducing their concentration by 66% and achieving a higher signal-to-background ratio than comparable spatially resolved immunostaining with static incubation.