Lab on a Chip最新文献

Nanoparticles have become widely used materials in various fields, yet their mechanism of action at the cellular level after entering the human body remains unclear. Accurately observing the effect of nanosize dimensions on particle internalization and toxicity in cells is crucial, particularly under the conditions of biological activity. With the aim of helping to study the interactions between nanoparticles of varying sizes and active cell membranes, we propose a flexible biosensor system based on a field effect transistor (FET). We constructed lipid bilayers on the device in vitro to simulate the interaction between nanoparticles and lipid membranes under active conditions, with the aim of investigating the effect of differently sized nanoparticles on the cell membrane. The experimental results revealed that nanoparticles with a diameter smaller than 50 nm tend to induce mild strain and repairable damage to the membrane, whereas nanoparticles larger than 50 nm may cause more severe damage, and even transmembrane penetration, by creating unrecoverable pores. The stretching of the lipid membrane exacerbated the deformation and destruction caused by nanoparticles, even in the case of smaller particles. These above results are consistent with previous theories on the interactions between cell membranes and nanoparticles. The proposed biosensors provide a valuable tool for investigating how the nanosize dimensions of particles affect their ability to penetrate and cause destruction in dynamic cell membranes, contributing to the improvement of a more comprehensive theoretical system for understanding the interaction process between nanoparticles and cell membranes.

Transepithelial electrical resistance (TEER) measurement is a label free, rapid and real-time technique, which is commonly used to evaluate the integrity of cell barriers. TEER characterization is important for applications, such as tissue (brain, intestines, lungs) barrier modeling, drug screening, and cell growth monitoring. Traditional TEER methods usually only show the average impedance of the whole cell layer, and lack accuracy and the characterization of internal spatial differences within cell layer regions. Here, we introduce a new spatial TEER strategy that utilizes microelectrode arrays (MEA) integrated in a Transwell to dynamically monitor TEER. A new electrical model which could reveal spatial impedance non-uniformity was proposed to extract accurate resistance from the measured data. Based on our method, the TEER signals from 16 different regions were successfully monitored in real time. The mapped impedance hotspots in different regions closely correlate with both fluorescence cell staining signals and calculated cell coverage, indicating the effectiveness of the developed spatial TEER system in monitoring local cell growth in vitro. The real-time spatial TEER responses to ethylene glycol-bis(β-aminoethylether)-N,N,N′,N′-tetraacetic acid (EGTA) and cisplatin were studied, which could either reduce barrier integrity or inhibit cellular growth. The obtained results demonstrated the spatial TEER's applicability for cell barrier function and cell growth monitoring. Our approach provides accurate spatial electrical information of cell barriers and holds potential applications in drug development and screening.

We present here a passive and label-free droplet microfluidic platform to sort cells stepwise by lactate and proton secretion from glycolysis. A technology developed in our lab, Sorting by Interfacial Tension (SIFT), sorts droplets containing single cells into two populations based on pH by using interfacial tension. Cellular glycolysis lowers the pH of droplets through proton secretion, enabling passive selection based on interfacial tension and hence single-cell glycolysis. The SIFT technique is expanded here by exploiting the dynamic droplet acidification from surfactant adsorption that leads to a concurrent increase in interfacial tension. This allows multiple microfabricated rails at different downstream positions to isolate cells with distinct glycolytic levels. The device is used to correlate sorted cells with three levels of glycolysis with a conventional surface marker for T-cell activation. As glycolysis is associated with both disease and cell state, this technology facilitates the sorting and analysis of crucial cell subpopulations for applications in oncology, immunology and immunotherapy.

Microfluidic-based sheath flow focusing methods have been widely used for efficiently isolating, concentrating, and detecting pathogenic bacteria for various biomedical applications due to their enhanced sensitivity and exceptional integration. However, such a microfluidic device usually needs complicated device fabrication and sample dilution, hampering the efficient and sensitive identification of target bacteria. In this study, we develop and fabricate a sheath-assisted and pneumatic-induced nano-sieve device for achieving the improved on-chip concentration and sensitive detection of Staphylococcus aureus (MRSA). The optimized nanochannel design with diverging configuration is beneficial to the regulation of the hydrodynamic flow while the sheath flow is focusing the sample to the confined region as expected. Per the experimental finding, a high flow ratio (sheath flow/sample flow) presents enhanced target concentration by comparing with a low flow ratio. With this setup, MRSA bacteria with an extremely low concentration of ∼100 CFU mL^-1 are successfully and sensitively detected under a fluorescence microscope, less than 30 min, demonstrating a reliable sheath-enhanced concentration and on-chip detection for target bacteria. Additionally, the theoretical model introduced here further rationalizes the working principle of our nano-sieve device, potentially guiding the optimization of next generation devices for highly sensitive and accurate on-chip bacteria detection at a much lower concentration level below 100 CFU mL^-1.

Modern cell and developmental biology increasingly relies on 3D cell culture systems such as organoids. However, routine interrogation with microscopy is often hindered by tedious, non-standardized sample mounting, limiting throughput. To address these bottlenecks, we have developed a pipeline for imaging intact organoids in flow, utilizing a transparent agarose fluidic chip that enables efficient and consistent recordings with theoretically unlimited throughput. The chip, cast from a custom-designed 3D-printed mold, is coupled to a mechanically controlled syringe pump for fast and precise sample positioning. We benchmarked this setup on a commercial digitally scanned light sheet microscope with cleared glioblastoma spheroids. Spheroids of varying sizes were positioned in the field of view with micrometer-level stability, achieving a throughput of 40 one-minute recordings per hour. We further showed that sample positioning could be automated through online feedback microscopy. The optical quality of the agarose chip outperformed FEP tubing, glass channels and PDMS casts for the clearing agents used, as demonstrated by image contrast profiles of spheroids stained with a fluorescent nuclear counterstain and further emphasized by the resolution of fine microglial ramifications within cerebral organoids. The retention of image quality throughout 500 μm-sized spheroids enabled comprehensive spatial mapping of live and dead cells based on their nuclear morphology. Finally, imaging a batch of LMNA knockout vs. wildtype astrocytoma spheroids revealed significant differences in their DNA damage response, underscoring the system's sensitivity and throughput. Overall, the fluidic chip design provides a cost-effective, accessible, and efficient solution for high-throughput organoid imaging.

Atherosclerosis is a chronic inflammatory vascular disorder driven by factors such as endothelial dysfunction, hypertension, hyperlipidemia, and arterial calcification, and is considered a leading global cause of death. Existing atherosclerosis models have limitations due to the absence of an appropriate hemodynamic microenvironment in vitro and interspecies differences in vivo. Here, we develop a simple but robust microfluidic intimal-lumen model of early atherosclerosis using interconnected dual channels for studying monocyte transmigration and foam cell formation at an arterial shear rate. To the best of our knowledge, this is the first study that creates a physiologically relevant microenvironment under an arterial shear rate to modulate lipid-laden foam cells on a microfluidic platform. As a proof of concept, we use murine endothelial cells to develop a vascular lumen in one channel and collagen-embedded murine smooth muscle cells to mimic the subendothelial intimal layer in another channel. The model successfully triggers endothelial dysfunction upon TNF-α stimulation, initiating monocyte adhesion to the endothelial monolayer under the arterial shear rate. Unlike existing in vitro models, native low-density lipoprotein (LDL) is added in the culture media instead of ox-LDL to stimulate subendothelial lipid accumulation, thereby mimicking more accurate physiology. The subendothelial transmigration of adherent monocytes and subsequent foam cell formation is also achieved under flow conditions in the model. The model also investigates the inhibitory effect of aspirin in monocyte adhesion and transmigration. The model exhibits a significant dose-dependent reduction in monocyte adhesion and transmigration upon aspirin treatment, making it an excellent tool for drug testing.

Osteoarthritis (OA) has long been considered a disease of the articular cartilage. Within the past decade it has become increasingly clear that OA is a disease of the entire joint space and that interactions between articular cartilage and subchondral bone likely play an important role in the disease. Driven by this knowledge, we have created a novel microphysiological model of the osteochondral unit containing synovium, cartilage, bone, and vasculature in separate compartments with molecular and direct cell–cell interaction between the cells from the different tissue types. We have characterized the model in terms of differentiation by molecule and matrix secretion and shown that it demonstrates morphology and functionality that mimic the native characteristic of the joint space. Finally, we induced inflammation and subsequently rescued the model constructs by a known compound as proof of concept for anti-inflammatory drug screening applications.

Microneedles hold the potential for enabling shallow skin penetration applications where biomarkers are extracted from the interstitial fluid (ISF) and drugs are injected in a painless and effective manner. To this purpose, needles must have an inner channel. Channeled needles were demonstrated using custom silicon microtechnology, having several needle tip geometries. Nevertheless, all the proposed fabrication sequences are not compatible with mass production based on mature, standard microfabrication techniques. Furthermore, ISF extraction was also demonstrated with channeled needles but under poorly controlled conditions and over long periods of time, the latter being impractical for medical use. A range of factors may impede or slow ISF extraction that require controlled experiments. In this work we address the above tasks in terms of microfabrication sequence design, tip geometry design and experimental validation under controlled conditions. We report the development and fabrication of a silicon channeled microneedle array using conventional, industrial micromechanic processes. With only 2 lithography steps, a hypodermic needle tip profile is achieved. Using the fabricated microneedles, fluid extraction is experimented on chicken skin mockups. Extraction tests are carried out by inducing a controlled pressure gradient between the two ends of the microneedle channels, generated by loading the chip or by applying vacuum to the chip's backside. The extraction of more than 1 μL of fluid in 20 minutes is demonstrated with a maximum applied pressure gradient of 500 mbar. A correlation between the extraction rate efficiency and needles' density is observed, both for short and long extraction times. These results provide the first demonstration of in vitro interstitial fluid collection under controlled experimental conditions using silicon hollow microneedles fabricated with standard micro electro mechanical systems (MEMS) fabrication technology and minimal steps. Based on the obtained data, a comparison is drawn between pressure load and vacuum as drivers for ISF extraction, according to modelling and controlled experiments.

The separation of large-size-range particles of complex biological samples is critical but yet well resolved. As a label-free technique, dielectrophoresis (DEP)-based particle separation faces the challenge of how to configure DEP in an integrated microfluidic device to bring particles of various sizes into the effective DEP force field. Herein, we propose a concept that combines the passive flow fraction mechanism with the accumulative DEP deflection effect in a cascaded manner. This concept places DEP deflection segments and bypass outlets alternately. Each DEP deflection segment is configured with an array of side-wall liquid metal electrodes to exert effective DEP forces on the particles of a suitable size range. After each DEP deflection segment, the passive bypass flow fraction mechanism diverts part of the sample flow and target range of particles through the bypass outlet. Simultaneously, this flow fraction brings the remaining particles closer to the electrodes in the subsequent DEP deflection segment, causing the next size range of particles to deflect under effective DEP forces and thus making them separable. Repeating this process, particles would be separated from the bypass outlets one by one in the order of reducing size ranges. We present the concept design and modeling, and prove the concept through separating five different particles ranging from 16–0.5 μm mixed together to mimic blood composition, providing a powerful platform for separating multiple particles in diverse biomedical applications.

Optoelectrowetting technology generates virtual electrodes to manipulate droplets by projecting optical patterns onto the photoconductive layer. This method avoids the complex design of the physical circuitry of dielectricwetting chips, compensating for the inability to reconstruct the electrode. However, the current technology relies on operators to manually position the droplets, draw optical patterns, and preset the droplet movement paths. It lacks real-time feedback on droplet information and the ability for independent droplet control, which can lead to droplet miscontrol and contamination. This paper presents a combination of optoelectrowetting with deep learning algorithms, integrating software and a photoelectric detection platform, and develops an optoelectrowetting intelligent control system. First, a target detection algorithm identifies droplet characteristics in real-time and automatically generate virtual electrodes to control movement. Simultaneously, a tracking algorithm outputs trajectories and ID information for efficient droplet arrays tracking. The results show that the system can automatically control the movement and fusion of multiple droplets in parallel and realize the automatic arrangement and storage of disordered droplet arrays without any additional electrodes and sensing devices. Additionally, through the automated control of the system, the cell suspension can be precisely cultured in the specified medium according to experimental requirements, and the growth trend is consistent with that observed in the well plate, significantly enhancing the experiment's flexibility and accuracy. In this paper, we propose an intelligent method applicable to the automated manipulation of discrete droplets. This method would play a crucial role in advancing the applications of digital microfluidic technology in biomedicine and other fields.