Review for "Direct access and recovery feature of solid precipitates embedded in microfluidic device"

Review for "Direct access and recovery feature of solid precipitates embedded in microfluidic device"

Author response for "Direct access and recovery feature of solid precipitates embedded in microfluidic device"

Review for "Direct access and recovery feature of solid precipitates embedded in microfluidic device"

Decision letter for "Direct access and recovery feature of solid precipitates embedded in microfluidic device"

Decision letter for "Direct access and recovery feature of solid precipitates embedded in microfluidic device"

Decision letter for "Direct access and recovery feature of solid precipitates embedded in microfluidic device"

Author response for "Direct access and recovery feature of solid precipitates embedded in microfluidic device"

Droplet microfluidics, which generates and manipulates water-in-oil microdroplets within continuous phases, has emerged as a compelling platform in modern science. The core advantage of this technology lies in the fact that each picoliter to nanoliter droplet functions as an independent microreactor, ensuring no cross-contamination. This enables ultra-high-throughput experiments while dramatically reducing the consumption of expensive reagents and rare samples. However, the efficient extraction of solid precipitates (such as crystals and particles) formed within droplets remains a fundamental challenge for subsequent analysis and utilization. This study proposes a novel microfluidic device and operational method to address these challenges: (1) the difficulty in extracting solids that cannot be recovered through simple fluid flow and (2) sample loss during long-distance transport. The key innovation combines (1) a passive trap structure for in situ solid formation processes within droplets and (2) a physically accessible harvesting chamber positioned nearby. This design eliminates the need for long-distance sample transport, enabling the gentle transfer of droplets containing precipitated solids to an adjacent extraction chamber with an open top, allowing for physical solid recovery. We demonstrated the system functionality using fluorescent microbeads as model particles, followed by the successful generation and recovery of protein (lysozyme) crystals as a practical application.

Review for "Direct access and recovery feature of solid precipitates embedded in microfluidic device"

Parkinson’s disease (PD) is the second major neurodegenerative disease and the most common movement disorder. Due to age being a critical risk factor, the rapid ageing of the world population further increases the prevalence of PD. So far no treatment is available and therapies mainly focus on motor symptoms by pharmacologically substituting striatal dopamine, caused by the loss of dopaminergic neurons in the substantia nigra. This neuronal loss and intracellular protein aggregates, termed Lewy bodies (LBs), are pathological characteristics of PD. With disease progression, a spread of LBs through the brain can be observed which mainly follows axonal projections. Understanding the mechanisms of this progressive spread could be central to discovering the underlying molecular pathogenesis of the disease. As LBs mainly consist of alpha-synuclein (-syn), a prion-like spreading of -syn was suggested and is now widely accepted as a component in the PD pathogenesis. New dopaminergic model systems to study the exact mechanisms underlying -syn spread are urgently needed. As PD is a human disease, in vitro models should be derived from humans. Lund human mesencephalic (LUHMES) cells are a suitable alternative to other, mostly non-human, dopaminergic cell lines. However, difficulties cultivating them in microfluidics devices has made them thus far inaccessible for co-cultivation studies in the field of PD spreading. In the first part of this thesis, a human dopaminergic cell model system for studying the spreading of -syn fibrils is presented. First, the well-characterized LUHMES cell line was tested for suitability of PD research on prion-like spreading, as no data is currently available on this matter. For the analysis, immunofluorescence light microscopy was employed. An extended period of differentiation aimed for a high degree of neuronal maturity and long neurites to facilitate the connectivity of spatially-separated cell populations. Seeding experiments with -syn fibrils revealed a weak toxicity against these assemblies, even at prolonged differentiation. Second, to study the transmission of -syn fibrils via neuronal projections, we developed a light microscopy-compatible microfluidic co-culturing device, to maintain two LUHMES cell populations in separate cell compartments for up to two weeks of differentiation. During this time, a neurite network is formed which connects the fluidically isolated cell growth compartments. The ability to cultivate cells with neurites and soma in an isolated environment enabled seeding and transmission experiments in anterograde and retrograde directions. In the second part of this thesis, implementation strategies of the microfluidic co-culturing chip for alternative analysis methods are discussed. Firstly, the accessibility of the cells in the co-culturing device using a single-cell lysis instrument is evaluated. The tool allows for targeted lysis of individual adherent cells. Preliminary tests point in a promising direction, while LUHMES single cell lysate was successfully transferred to different analysis techniques. However, direct access to the channels of the microfluidic co-culturing chip was problematic and needs further modifications. Secondly, an implementation of the microfluidic device aiming for co-cultivation of LUHMES cells on electron microscopy grids to study neurite architecture was pursued. Thereby, microfluidic devices harbor only cell soma, but neurites can grow onto an electron microscopy grid, as only they are thin enough to be visualized by cryo-electron microscopy. Proof-of-concept experiments demonstrate the direct visualization of LUHMES cell neurites in a near-native, frozen-hydrated state.

We introduce a novel microfluidic device to co-culture a blood vessel network and cell tissues in an in vivo-like niche. Our "open-top" microfluidic device is composed of microchannels with micropores in the ceiling, which provides direct fluid access from reservoir to microchannel. Fluid connections through micropores afford novel advantages, including: i) the long-term culture of large-scale microvessel network, ii) access of different fluids to inner and exterior sides of the microvessel, and iii) co-culturing of the microvessel network and small cell tissue. In this study, we have successfully assembled microvessels with 5 mm channel widths. We were also able to mimic capillary bed conditions by co-culturing microvessels with cancer spheroids. Intimate contact between the cancer spheroid and microvessel caused vessel recruitment and an increase in vessel formation, and affected vessel morphology. We expect this device to be used as a novel platform for vascularized tissue models.

By utilizing the coffee-ring effect and microfluidic paper-based analytical devices (µPADs), this study improved the sensitivity of the determination of norfloxacin in four different food matrices. Micro-PADs in this study were fabricated by designing and embedding wax channels onto cellulose-based filter paper through printing and subjecting the paper to heat to allow the wax to penetrate the paper. Determination of norfloxacin concentration in food samples was achieved by monitoring the colorimetric reaction that occurred between norfloxacin and the added iron (III) nitrate nonahydrate in 5mM ammonia in each reaction chamber. A transition metal hydroxide was formed through this reaction that resulted in the formation of a solid precipitate to enable the antibiotic to bind to the iron molecule via coordination chemistry. This metal ion-antibiotic complex generated a visible color change. Following the colorimetric reaction, images were taken and subsequently analyzed via ImageJ to determine the relative pixel intensity that was used to infer norfloxacin concentration. The analytical sensitivity of this device was determined to be as low as 50ppm when analyzing the inner-ring reaction, and as low as 5ppm when analyzing the outer coffee ring thereby allowing for an alternative cheaper, faster, and more user-friendly method to detect norfloxacin than the conventional methods. PRACTICAL APPLICATION: This novel paper-based microfluidic device can achieve the detection of antibiotic residues in agrifoods in a faster, cheaper, and more user-friendly manner.

Techniques, such as micropipette aspiration and optical tweezers, are widely used to measure cell mechanical properties, but are generally labor-intensive and time-consuming, typically involving a difficult process of manipulation. In the past two decades, a large number of microfluidic devices have been developed due to the advantages they offer over other techniques, including transparency for direct optical access, lower cost, reduced space and labor, precise control, and easy manipulation of a small volume of blood samples. This review presents recent advances in the development of microfluidic devices to evaluate the mechanical response of individual red blood cells (RBCs) and microbubbles flowing in constriction microchannels. Visualizations and measurements of the deformation of RBCs flowing through hyperbolic, smooth, and sudden-contraction microchannels were evaluated and compared. In particular, we show the potential of using hyperbolic-shaped microchannels to precisely control and assess small changes in RBC deformability in both physiological and pathological situations. Moreover, deformations of air microbubbles and droplets flowing through a microfluidic constriction were also compared with RBCs deformability.

Release mechanisms of PLGA microparticles prepared using a microfluidics device or a beaker

With the extensive application of the Direct Memory Access (DMA) technique, the efficiency of data transfer between the peripheral and the host machine has been improved dramatically. However, these optimizations also introduce security vulnerabilities and expose the process of data transmission to DMA attacks that utilize the feature of direct access to steal the data stored in the live memory on the victim system. In this paper, we propose a lightweight scheme to provide resilience to DMA attacks without physical and protocol-level modification. The proposed scheme constructs a unique identifier for each DMA-supported PCIe device based on profiling time and builds a trusted database for authentication. The experimental result shows that the proposed methodology eliminates most of the noise produced in the measuring process for identifier construction and the success rate of authentication is 100% for all the devices.