The COVID-19 pandemic has underscored the emergent need for efficient and scalable methods to predict viral outbreaks at the community level. Wastewater-based epidemiology (WBE) has proven effective in tracking the prevalence of COVID-19 infection, but conventional methods rely heavily on sophisticated laboratory infrastructure and trained technical personnel, limiting their accessibility in resource-constrained settings. This study presents a simplified wastewater processing approach that integrates ultrafiltration with a dipstick-based RNA extraction method to enable rapid, low-cost preparation of samples for downstream detection by reverse transcription quantitative polymerase chain reaction (RT-qPCR). Ultrafiltration with a sequential filter system effectively concentrates viral particles, while dipstick extraction provides a fast, equipment-minimal RNA purification step. Using SARS-CoV-2-spiked wastewater samples, the simplified workflow produced C-t values comparable to those obtained with conventional laboratory methods. By substantially reducing processing time, reagent cost, and dependence on specialized equipment, this approach addresses major barriers to broad WBE implementation. This makes it particularly well-suited for deployment in decentralized or resource-limited settings. Future work will focus on integrating this simplified approach with a portable PCR platform for point-of-care virus detection, facilitating timely public health interventions, and supporting global pandemic preparedness.

Pathological blood clots cause life-threatening events, including ischemic stroke and myocardial infarction. Tissue plasminogen activator (TPA), a clinically approved fibrinolytic agent, remains a frontline therapy for clot dissolution. However, its short plasma half-life and requirement for high systemic doses pose serious risks, particularly hemorrhagic complications. To address these limitations, a shear-responsive delivery platform was engineered for targeted TPA delivery at sites of pathological high shear. This system, termed Single-MOlecule–based materials with structures and functions REsponsive to Shear (SMORES), consists of a shear-amplifying microparticle conjugated to a TPA-binding aptamer and functionalized with a monoclonal antibody against integrin αIIbβ3 to selectively target activated platelets within thrombi. Under elevated shear in stenotic vessels, the immobilized aptamer undergoes mechanical unfolding, triggering localized TPA release. Optical tweezers–based single-molecule force measurements probing aptamer unfolding and refolding under constant force and varied relaxation periods revealed a narrow range of threshold forces (4–10 pN) governing the transition between folded and unfolded states. Under defined flow conditions in a microfluidic channel, single-molecule flow experiments demonstrated a peak in TPA release at 50 μL min−1. COMSOL-based fluid dynamics simulations estimated a corresponding tether force of 4.76 pN and a shear rate of 3,512.9 s−1, consistent with pathological shear levels observed in stenotic vessels and aligning with optical tweezers predictions. The enzymatic activity of released TPA was confirmed by a clot dissolution assay, demonstrating maximal fibrin degradation under the same flow condition. Together, these results demonstrate that TPA-SMORES enables mechanistically controlled, site-specific thrombolysis, offering a promising strategy to improve thrombolytic precision.

Patients with continuous-flow ventricular assist devices (CF-VADs) face an elevated risk of nonsurgical bleeding. One hypothetical cause is that the loss of pulsatility promotes unraveling and enzymatic degradation of von Willebrand factor (VWF), a key clotting protein. Artificial pulsatility has been proposed to counter this effect, but the role of pulse frequency in VWF unraveling remains unclear. This study investigates VWF conformational changes in response to varying pulse frequencies. Membrane-bound VWF on human aortic endothelial cells (HAECs) exposed to pulsatile in vitro conditions exhibited significantly less unraveling than under continuous flow ( p < 0.005). To enable real-time observation of VWF conformation, VWF was immobilized in a microfluidic device and exposed to continuous or pulsatile flows (20, 40, or 60 pulses/min) to model HAEC-bound unraveling. Results showed that frequencies greater than or equal to 40 pulses/min significantly reduced maximum extension compared with continuous flow and low-frequency conditions (≤ 20 pulses/min), whereas minimum extension was greatest under continuous flow and declined as frequency increased. Step-change flow experiments revealed a time constant of 0.19 ± 0.04 seconds for extension and ~1 second for recoiling. These findings support optimizing pulsatile flow frequency as a strategy to minimize VWF unfolding and mitigate nonsurgical bleeding in CF-VAD patients.

Thermophoresis, the directed motion of particles along a temperature gradient, is a technique used for separating molecules and colloidal particles. However, few methods exist that can simultaneously measure local fluid properties and thermophoretic motion. This hinders the optimization of thermophoretic separation in complex fluids, where local rheology is temperature-dependent. We use two-dimensional (2D) multiple particle tracking microrheology with a 1D temperature gradient to measure thermal-gradient driven motion in the x-direction and temperature-dependent viscosity from Brownian motion in the y-direction. We measure viscosity and particle motion in Newtonian glycerol-water solutions in ground (0-50 w/w%) and microgravity conditions (20 and 30 w/w%) on the International Space Station (ISS) and compare our results to tabulated data. Microgravity eliminates buoyancy-induced recirculation, enabling measurement of only thermophoretic motion. Viscosities measured in the y-direction in ground and microgravity conditions agree with tabulated values. In ground conditions, measured particle velocities in the x-direction scale with viscosity and thermal gradient, consistent with recirculation. In microgravity, particle motion is Brownian in both directions. Although the apparent Soret coefficient values in microgravity indicate thermophoretic motion, the absence of directed motion and no measurable difference between motion in the x- and y-directions show that thermophoresis is undetectable. This is likely due to an insufficient temperature gradient achievable in our experiments on the ISS. Our method enables measurements of local viscosity orthogonally to an applied temperature gradient and can be used to measure spatial variations in local rheology, which is essential for quantifying thermal-gradient driven motion in complex fluids.

The dielectric property of biological cells exhibits a rich frequency-dependent behavior that is related to cellular structure, function, and molecular makeup. Assessing the dielectric properties of single cells offers a label-free and noninvasive method for disease diagnosis, cell identification, and monitoring of cellular structure and function. This article reviews the fundamentals and applications of microwave biosensors for single-cell analysis, with a focus on how electromagnetic interactions across the and dispersion regimes can be leveraged to extract cellular parameters such as membrane capacitance, cytoplasmic conductivity, and intracellular hydration. We begin by discussing the physical principles of dielectric dispersion and the equivalent circuit models in biological cells that underpin microwave-based sensing. We then examine major classes of sensing platforms, including transmission-line sensors, capacitive electrode arrays, and resonant structures, highlighting their design strategies, strength and weakness, and applications in single-cell sensing. The role of dielectrophoresis (DEP) as both a manipulation tool and sensing mechanism is also discussed, particularly in the context of hybrid systems that combine low-frequency trapping with high-frequency dielectric readout. Together, these technologies represent a convergence of microwave engineering, biophysics, and microsystem design to enable high-resolution, real-time interrogation of single cells. After describing the theoretical foundations with recent experimental advances, we provide a perspective on the design of next-generation dielectric biosensors tailored to the demands of single-cell diagnostics, drug screening, and functional phenotyping.

Non-surgical bleeding is a common complication in patients on continuous flow left ventricular assist device (CF-VAD) support. This study investigates how the transition from cyclic to constant stretch following CF-VAD implantation affects endothelial biosynthesis and release of Von Willebrand factor (VWF) and angiopoietin-2 (ANGPT-2), two molecules that play an essential role in the development of non-surgical bleeding. Human aortic endothelial and umbilical vein endothelial cells (HAECs and HUVECs) were cultured within a uniaxial stretch device that mimics stretch associated with both normal pulsatile and CF-VAD conditions. Following 72 hours of stretch, transcriptional regulation, intracellular accumulation, and secretion of VWF and ANGPT-2 were evaluated using molecular expression profiling and immunofluorescence microscopy. Constant stretch associated with CF-VADs upregulates transcriptional levels of VWF and ANGPT-2 in HAECs and HUVECs compared to physiological cyclic stretch (p < 0.05). Transcriptional increases in both VWF and ANGPT-2 in HAECs also resulted in increased intracellular protein levels of VWF and ANGPT-2 measured using ELISA, western blots and immunofluorescence microscopy, whereas in HUVECs, the intracellular increase was evident only with western blots and immunofluorescence microscopy. Finally, constant stretch appears to promote ANGPT-2 release and inhibit release of VWF from both HAECs and HUVECs compared to cyclic stretch. Our study found that constant stretch upregulates the production of both VWF and ANGPT-2. However, while the release of ANGPT-2 is elevated under constant stretch, the release of VWF declines, resulting in elevated extracellular levels of ANGPT-2, but not VWF.

Background: Continuous Flow Left Ventricular Assist Devices (CF-LVADs) are the gold standard therapy for heart failure (HF) patients. Although this therapeutic approach is associated with improved clinical outcomes, CF-LVAD patients experience non-physiological reductions in arterial pressure and flow pulsatility, which are linked to an increased risk of nonsurgical bleeding, and arteriovenous malformations (AVMs). Studies suggest reduced pulsatility leads to von Willebrand factor (VWF) degradation and Angiopoietin-2 (ANGPT-2) induction in the bloodstream. While their interaction in endothelial cells is observed, their combined effects in CF-LVAD patients are underexplored. This study evaluates the cooperative effect of VWF and ANGPT-2 on non-surgical bleeding in CF-LVAD patients. Methods: First, we quantitatively assessed VWF and ANGPT-2 levels in the blood of CF-LVAD patients (n=19), both before VAD implantation and monthly thereafter, using ELISA and Western blotting. The impact of pulsatility loss on VWF/ANGPT-2 status and angiogenesis was assessed using an in vitro vascular pulse perfusion model (VPPM). Plasma samples from CF-LVAD patients were analyzed for VWF and ANGPT-2 complexes via immunoprecipitation. Additionally, endothelial cells were treated with recombinant ANGPT-2 and VWF to assess angiogenic marker expression. Results: High molecular weight (HMW) VWF levels significantly decreased in all CF-LVAD patients. In contrast, ANGPT-2 levels transiently increased in patients experiencing bleeding, whereas LVAD patients without bleeding showed decreased ANGPT-2 levels. Our results suggest that increased ANGPT-2 and the loss of HMW-VWF may predict bleeding in CF-LVAD patients. Using VPPM, we mimicked the stretch generated during CF-LVAD flow and observed induced angiogenesis, with marked upregulation of VWF and ANGPT-2. Next, we evaluated the cooperative promotion of angiogenesis by VWF and ANGPT-2. Immunoprecipitation confirmed the presence of VWF and ANGPT-2 complexes in patients’ plasma. Treatment of endothelial cells with recombinant ANGPT-2 and VWF proteins significantly increased the expression of angiogenic markers at both the transcriptional and translational levels. Conclusion: Our data reveal that VWF and ANGPT-2 work cooperatively in developing non-surgical bleeding, potentially by inducing angiogenesis, and can serve as biomarkers for predicting bleeding risk and guiding treatment in CF-LVAD patients.

Sickle cell disease (SCD) poses a significant yet under-addressed public health challenge in Sierra Leone where limited diagnostic infrastructure, insufficient medical training, and a lack of unified national protocols hinder effective care and management [1]. The SicklED team at Lehigh University presents a comprehensive five-point strategy aimed at transforming SCD outcomes in the region through accessible, sustainable, and context-specific interventions. This paper outlines the initiative for the development of a low-cost, point-of-care screening tool to increase early detection; the implementation of targeted medical education initiatives to bolster provider knowledge; the distribution of culturally adapted educational materials to enhance community awareness; the creation of nationally recognized supplemental guidelines and a standard operating procedure (SOP) to unify SCD care protocols; and the launch of a robust data collection framework to support evidence-based policy and health system planning. Through interdisciplinary collaboration, global partnerships, and ongoing feedback from Sierra Leonean healthcare professionals and advocacy groups, the SicklED strategy aims to catalyze systemic change and provide a replicable model for addressing noncommunicable diseases in resource-limited settings.

Smart polymers that mimic and even surpass the functionality of natural responsive materials have been actively researched. This study explores the design and characterization of a Single-MOlecule-based material REsponsive to Shear (SMORES) for the targeted release of A1, the platelet binding domain of the blood clotting protein von Willebrand factor (VWF). Each SMORES construct employs an aptamer molecule as the flow transducer and a microparticle to sense and amplify the hydrodynamic force. Within the construct, the aptamer, ARC1172, undergoes conformational changes beyond a shear stress threshold, mimicking the shear-responsive behavior of VWF. This conformational alteration modulates the bioavailability of its target, the VWF-A1 domain, ultimately releasing it at elevated shear. Through optical tweezer-based single-molecule force measurement, ARC1172s role as a force transducer was assessed by examining its unfolding under constant pulling force. We also investigated its refolding rate as a function of force under varied relaxation periods. These analyses revealed a narrow range of threshold forces (3-7 pN) governing the transition between folded and unfolded states. We subsequently constructed the SMORES material by conjugating ARC1172 and a microbead, and immobilizing the other end of the aptamer on a substrate. Single-molecule flow experiments on immobilized SMORES constructs revealed a peak A1 domain release within a flow rate range of (40-70 mu L min-1). A COMSOL Multiphysics model translated these flow rates to total forces of 3.10 pN-5.63 pN experienced by the aptamers, aligning with single-molecule force microscopy predictions. Evaluation under variable flow conditions showed a peak binding of A1 to the platelet glycoprotein Ib (GPIB) within the same force range, confirming released payload functionality. Building on knowledge of aptamer biomechanics, this study presents a new strategy to create shear-stimulated biomaterials based on single biomolecules. (A). Optical tweezers characterized the mechanical properties of the Single-MOlecule-based material REsponsive to Shear (SMORES). (B) Flow experiments demonstrated the release of therapeutic cargo at a threshold flow rate.

Myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) is a demanding medical condition for patients and society. It has raised much more public awareness after the COVID-19 pandemic since ME/CFS and long-COVID patients share many clinical symptoms such as debilitating chronic fatigue. However, unlike long COVID, the etiopathology of ME/CFS remains a mystery despite several decades' research. This review moves from pathophysiology of ME/CFS through the compelling evidence and most interesting hypotheses. It focuses on the pathophysiology of skeletal muscle by proposing the hypothesis that skeletal muscle tissue offers novel opportunities for diagnosis and treatment of this syndrome and that new evidence can help resolve the long-standing debate on terminology.

Three-dimensional computational fluid dynamics simulations were run to simulate the nanoparticle concentrating in thermophoresis microfluidic devices. Thermophoresis is a phenomenon where particles are driven by a temperature gradient. The microfluidic device utilizes thermophoretic force by applying 10 K temperature difference across the channel. The artificial transverse flow induced by slanted grooves is combined with the thermophoresis to concentrate nanoparticles. Following the experimental research, this study revisited the theory by applying the concept of circulation to evaluate the intensity of the transverse flow. Apart from the original experimental setup with 175 mu m groove spacing, 25 mu m and 75 mu m setups were also simulated as comparison. The comparison shows that the peak circulation has no correlation between different groove spacing. The mass concentration accumulation was analyzed, and smaller spacing resulted in less fluctuation and more effective particle focusing. The averaged mass concentration from the outlet was processed, and in the best scenario, the 25 mu m spacing design achieved 68% more concentration than the original 175 mu m spacing design from the experiment.

Nonsurgical bleeding occurs in a significant proportion of patients implanted with continuous-flow ventricular assist devices (CF-VADs) and is associated with nonphysiologic flow with diminished pulsatility. An in vitro vascular pulse perfusion model seeded with adult human aortic endothelial cells (HAECs) was used to identify biomarkers sensitive to changes in pulsatility. Diminished pulsatility resulted in an ~45% decrease in von Willebrand factor (vWF) levels from 9.80 to 5.32 ng/ml (n = 5, p < 0.05) and a threefold increase in angiopoietin-2 (ANGPT-2) levels from 775.29 to 2471.93 pg/ml (n = 5, p < 0.05) in cultured HAECs. These changes are in agreement with evaluation of patient blood samples obtained pre-CF-VAD implant and 30-day postimplant: a decrease in plasma vWF level by 50% from ~45.59 to ~22.49 μg/ml (n = 15, p < 0.01) and a 64% increase in plasma ANGPT-2 level from 7,073 to 11,615 pg/ml (n = 8, p < 0.05). This study identified vWF and ANGPT-2 as highly sensitive to changes in pulsatility, in addition to interleukin-6 (IL-6), IL-8, and tumor necrosis-α (TNF-α). These biomarkers may help determine the optimal level of pulsatility and help identify patients at high risk of nonsurgical bleeding.

Many skeletal muscle diseases such as muscular dystrophy, myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS), and sarcopenia share the dysregulation of calcium (Ca2+) as a key mechanism of disease at a cellular level. Cytosolic concentrations of Ca2+ can signal dysregulation in organelles including the mitochondria, nucleus, and sarcoplasmic reticulum in skeletal muscle. In this work, a treatment is applied to mimic the Ca2+ increase associated with these atrophy-related disease states, and broadband impedance measurements are taken for single cells with and without this treatment using a microfluidic device. The resulting impedance measurements are fitted using a single-shell circuit simulation to show calculated electrical dielectric property contributions based on these Ca2+ changes. From this, similar distributions were seen in the Ca2+ from fluorescence measurements and the distribution of the S-parameter at a single frequency, identifying Ca2+ as the main contributor to the electrical differences being identified. Extracted dielectric parameters also showed different distribution patterns between the untreated and ionomycin-treated groups; however, the overall electrical parameters suggest the impact of Ca2+-induced changes at a wider range of frequencies.

Individual cells have many unique properties that can be quantified to develop a holistic understanding of a population. This can include understanding population characteristics, identifying subpopulations, or elucidating outlier characteristics that may be indicators of disease. Electrical impedance measurements are rapid and label-free for the monitoring of single cells and generate large datasets of many cells at single or multiple frequencies. To increase the accuracy and sensitivity of measurements and define the relationships between impedance and biological features, many electrical measurement systems have incorporated machine learning (ML) paradigms for control and analysis. Considering the difficulty capturing complex relationships using traditional modelling and statistical methods due to population heterogeneity, ML offers an exciting approach to the systemic collection and analysis of electrical properties in a data-driven way. In this work, we discuss incorporation of ML to improve the field of electrical single cell analysis by addressing the design challenges to manipulate single cells and sophisticated analysis of electrical properties that distinguish cellular changes. Looking forward, we emphasize the opportunity to build on integrated systems to address common challenges in data quality and generalizability to save time and resources at every step in electrical measurement of single cells.

Thermal diffusion of particles in dilute aqueous suspensions is driven by the interactions between the dispersing medium and the particle, which are largely influenced by the properties of the medium. Using a commercial instrument to generate thermophoresis, we developed a method to quantify the migration of colloids in a temperature gradient and further studied how it varies based on the composition and pH of the dispersing medium and with an anionic surfactant, at different salt concentrations. Thermophoretic migration of aqueous suspensions of carboxylate-modified polystyrene particles with different compositions is measured as MicroScale Thermophoresis (MST) traces and a mathematical model is developed to extract the Soret coefficient (ST). Soret coefficient measurements obtained using the developed method are in-line with previous theories and scientific findings from other literature, indicating a dependence of the ST on the Debye length and surface charge density of the suspended particles, both of which are controlled by the composition of the dispersing medium. The thermophobic/thermophilic behavior of particles is also found to be strongly influenced by the thermoelectric effect of the buffer ions. In this paper, a new analytical model is introduced and applied to complex systems to understand their thermophoretic behavior as a function of solvent properties.

Many recent efforts in the diagnostic field address the accessibility of cancer diagnosis. Typical histological staining methods identify cancer cells visually by a larger nucleus with more condensed chromatin. Machine learning (ML) has been incorporated into image analysis for improving this process. Recently, impedance spectrometers have been shown to generate all-inclusive lab-on-a-chip platforms to detect nucleus abnormities. In this paper, a wideband electrical sensor and data analysis paradigm that can identify nuclear changes shows the realization of a single-cell microfluidic device to detect nuclei of altered sizes. To model cells of altered nucleus, Jurkat cells were treated to enlarge or shrink their nucleus followed by broadband sensing to obtain the S-parameters of single cells. The ability to deduce important frequencies associated with nucleus size is demonstrated and used to improve classification models in both binary and multiclass scenarios, despite a heterogeneous and overlapping cell population. The important frequency features match those predicted in a double-shell circuit model published in prior work, demonstrating a coherent new analytical technique for electrical data analysis. The electrical sensing platform assisted by ML with impressive accuracy of cell classification looks forward to a label-free and flexible approach to cancer diagnosis.

The COVID-19 pandemic has presented a significant challenge to the world's public health and led to over 6.9 million deaths reported to date. A rapid, sensitive, and cost-effective point-of-care virus detection device is essential for the control and surveillance of the contagious severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) pandemic. The study presented here aimed to demonstrate a solid-phase isothermal recombinase polymerase amplification coupled CRISPR-based (spRPA-CRISPR) assay for on-chip multiplexed, sensitive and visual COVID-19 DNA detection. The assay targets the SARS-CoV-2 structure protein encoded genomes and can simultaneously detect two specific genes without cross-interaction. The amplified target sequences were immobilized on the one-pot device surface and detected using the mixed Cas12a-crRNA collateral cleavage of reporter-released fluorescent signal when specific genes were recognized. The endpoint signal can be directly visualized for rapid detection of COVID-19. The system was tested with samples of a broad range of concentrations (20 to 2 × 104 copies) and showed analytical sensitivity down to 20 copies per microliter. Furthermore, a low-cost blue LED flashlight (∼$12) was used to provide a visible SARS-CoV-2 detection signal of the spRPA-CRISPR assay which could be purchased online easily. Thus, our platform provides a sensitive and easy-to-read multiplexed gene detection method that can specifically identify low concentration genes.

Background: Non-surgical bleeding is one of the most common adverse events associated with continuous flow (CF) ventricular assist devices (VADs), which is linked to shear mediated degradation of von Willebrand factor (vWF). vWF degradation has traditionally been studied in the context of supraphysiological shear stresses within the VAD. However, recent evidence suggests that loss of pulsatility is a significant contributing factor for vWF degradation and in vitro systems are needed to study this mechanism. Methods: A compact, pneumatically driven, in vitro human arterial endothelial cell culture model, that incorporates directional control, resistance, compliance, and pressure and flow measurements was developed (Fig. 1). The device has a microfluidic channel for seeding human arterial endothelial cells (HAECs). It was cast in PDMS from a mold and plasma bonded to a glass cover slip to permit imaging. Flow and pressure data were collected using custom LabView scripts. Results: Our model was able to simulate physiological pulse pressure, flow profile, and shear rates for normal pulsatile flow (Fig 2A). By altering the compliance element volumes, we also simulated the flow, pressure, and shear profiles experienced with CF-VAD support (Fig 2B). HAECs were successfully cultured in the microfluidic channel and imaged under microscopy (Fig 3). Conclusion: By being able to modulate flow, shear, and pressure magnitude and amplitude, our model system will enable the study of how pressure and flow characteristics affect vWF secretion, unraveling, and degradation. These data may inform changes to VAD flow modulation or VAD design to minimize bleeding. Figure 1: In vitro perfusion loop model Figure 2: Pressure, flow, and shear profiles produced to simulate (A) normal/pulsatile and (B) CF-VAD supported/continuous flow conditions Figure 3: (A) Phase contrast and (B) DAPI microscopy images of Human aortic endothelial cells (HAECs) cultured in the microfluidic channel

The advances of biomedicine and biotechnology demand new approaches to enrich biological nanoparticles, such as viruses, viral vectors and nanovesicles, in an easy-to-operate fashion. Conventional methods, such as ultracentrifugation and ultrafiltration, require bulky instruments and extensive manual operation. Inspired by recent research of thermophoresis of biomolecules and bio-nanoparticles in aqueous solutions, we present a microfluidic design that directly focuses nanoparticles in a label-free and flow-through process by coupling an engineered swirling flow and a moderate, one-dimensional temperature gradient. Enrichment of polystyrene particles, HIV and bacteriophage samples was quantitatively determined, indicating the compatibility of the microfluidic approach with synthetic and biological samples. The focusing results are well predicted using a numerical model. As thermophoresis is ubiquitous, the microfluidic approach can be applied broadly to bio-nanoparticle enrichment without the necessity of labeling, buffer exchange, or sheath fluids, permitting continuous retrieval of concentrated species in a simple, controlled flow with little infrastructure needs.