Microfluidic Features Research Articles

Stacked hydrogel-based brain-on-chips utilizing capillary force flow pinning

Cellular processes in brain tissue such as migration, proliferation, morphology, and differentiation are influenced by mechanical cues, demonstrating the interplay between the structure and function. Given the complexity, it remains a substantial challenge to establish a reliable in vitro model mimicking structural properties as brain tissue. To address this challenge, we propose an innovative approach to create vertical hydrogel stacks based on microfluidic technology. 3D-printed microfluidic features in the sidewall profile of our chip designs allowed us to faithfully replicate these capillary force flow pinning structures in polydimethylsiloxane. After the successful application of hydrogel of a defined height, thanks to the pinning process and plating of stem-cell-derived neurons, the results demonstrated the potential of our BoC platform, providing a valuable tool for neuroscience research.

Advantages of stereolithographic 3D printing in the fabrication of the Affiblot device for dot-blot assays

In stereolithographic (SLA) 3D printing, objects are constructed by exposing layers of photocurable resin to UV light. It is a highly user-friendly fabrication method that opens a possibility for technology sharing through CAD file online libraries. Here, we present a prototyping procedure of a microfluidics-enhanced dot-blot device (Affiblot) designed for simple and inexpensive screening of affinity molecule characteristics (antibodies, oligonucleotides, cell receptors, etc.). The incorporation of microfluidic features makes sample processing user-friendly, less time-consuming, and less laborious, all performed completely on-device, distinguishing it from other dot-blot devices. Initially, the Affiblot device was fabricated using CNC machining, which required significant investment in manual post-processing and resulted in low reproducibility. Utilization of SLA 3D printing reduced the amount of manual post-processing, which significantly streamlined the prototyping process. Moreover, it enabled the fabrication of previously impossible features, including internal fluidic channels. While 3D printing of sub-millimeter microchannels usually requires custom-built printers, we were able to fabricate microfluidic features on a readily available commercial printer. Open microchannels in the size range 200–300 μm could be fabricated with reliable repeatability and sealed with a replaceable foil. Economic aspects of device fabrication are also discussed.Graphical abstract

Droplet-Based Microfluidics Reveals Insights into Cross-Coupling Mechanisms over Single-Atom Heterogeneous Catalysts.

Single-atom heterogeneous catalysts (SACs) hold promise as sustainable alternatives to metal complexes in organic transformations. However, their working structure and dynamics remain poorly understood, hindering advances in their design. Exploiting the unique features of droplet-based microfluidics, we present the first in-situ assessment of a palladium SAC based on exfoliated carbon nitride in Suzuki-Miyaura cross-coupling using X-ray absorption spectroscopy. Our results confirm a surface-catalyzed mechanism, revealing the distinct electronic structure of active Pd centers compared to homogeneous systems, and providing insights into the stabilizing role of ligands and bases. This study establishes a valuable framework for advancing mechanistic understanding of organic syntheses catalyzed by SACs.

MscI restriction enzyme cooperating recombinase-aided isothermal amplification for the ultrasensitive and rapid detection of low-abundance EGFR mutations on microfluidic chip

The detection of low-abundance mutation genes of the epidermal growth factor receptor (EGFR) exon 21 (EGFR L858R) plays a crucial role in the diagnosis of non-small cell lung cancer (NSCLC), as it enables early cancer detection and facilitates the development of treatment strategies. A detection platform was developed by combining the MscI restriction enzyme with the recombinase-aided isothermal amplification (RAA) technique (MRE-RAA). During the RAA process, “TGG^CCA” site of the wild-type genes was cleaved by the MscI restriction enzyme, while only the low-abundance mutation genes underwent amplification. Notably, when the RAA product was combined with CRISPR-Cas system, the sensitivity of detecting the EGFR L858R mutation increased by up to 1000-fold for addition of the MscI restriction enzyme. This achievement marked the first instance of attaining an analytical sensitivity of 0.001%. Furthermore, a disk-shaped microfluidic chip was developed to automate pretreatment while concurrently analyzing four blood samples. The microfluidic features of the chip include DNA extraction, MRE-RAA, and CRISPR-based detection. The fluorescence signal is employed for detection in the microfluidic chip, which is visible to the naked eye upon exposure to blue light irradiation. Furthermore, this platform has the capability to facilitate early diagnosis for various types of cancer by enabling high-sensitivity detection of low-abundance mutation genes.

A Rapid Prototyping Approach for Multi-Material, Reversibly Sealed Microfluidics.

Microfluidic organ-on-chip models recapitulate increasingly complex physiological phenomena to study tissue development and disease mechanisms, where there is a growing interest in retrieving delicate biological structures from these devices for downstream analysis. Standard bonding techniques, however, often utilize irreversible sealing, making sample retrieval unfeasible or necessitating destructive methods for disassembly. To address this, several commercial devices employ reversible sealing techniques, though integrating these techniques into early-stage prototyping workflows is often ignored because of the variation and complexity of microfluidic designs. Here, we demonstrate the concerted use of rapid prototyping techniques, including 3D printing and laser cutting, to produce multi-material microfluidic devices that can be reversibly sealed. This is enhanced via the incorporation of acrylic components directly into polydimethylsiloxane channel layers to enhance stability, sealing, and handling. These acrylic components act as a rigid surface separating the multiple mechanical seals created between the bottom substrate, the microfluidic features in the device, and the fluidic interconnect to external tubing, allowing for greater design flexibility. We demonstrate that these devices can be produced reproducibly outside of a cleanroom environment and that they can withstand ~1 bar pressures that are appropriate for a wide range of biological applications. By presenting an accessible and low-cost method, we hope to enable microfluidic prototyping for a broad range of biomedical research applications.

Inexpensive and rapid fabrication of PDMS microfluidic devices for biological testing applications using low cost commercially available 3D printers

Polydimethylsiloxane (PDMS) elastomers have been extensively used in the development of microfluidic devices, capable of miniaturizing biomolecular and cellular assays to the microlitre and nanolitre range, thereby increasing the throughput of experimentation. PDMS has been widely used due to its optical clarity and biocompatibility, among other desirable physical and chemical properties. Despite the widespread use of PDMS in microfluidic devices, the fabrication process typically via soft lithography technology requires specialized facilities, instruments, and materials only available in a limited number of laboratories. To expand microfluidic research capabilities to a greater scientific population, we developed and characterized a simple and robust method of fabricating relatively inexpensive PDMS microfluidic devices using readily available reagents and commercially available three-dimensional (3D) printers. The moulds produced from the 3D printers resolve designed microfluidic channel features accurately with high resolution (>100 µm). The critical physical and chemical post-processing modifications we outline here are required to generate functional and optically clear microfluidic devices.

Development of microfluidic chip flowmeter-based constant pressure system for analysing the hydrogen adsorption performance of non-evaporable getters

BackgroundAs one of the primary residual gases in vacuum, hydrogen affects the performance of MEMS devices. It commonly uses a non-evaporable getter (NEG) to adsorb hydrogen in this case. One of the standard test methods for NEG is the constant pressure method. However, most constant pressure test systems control the intake flow by valves or small orifices. These methods are crude and limit the reliability of the result. Therefore, it is necessary to provide a stable intake flow method for the constant pressure test system to improve the accuracy of the test. ResultsWe demonstrate a constant pressure system based on the microfluidic chip flowmeter to evaluate the hydrogen adsorption performance of non-evaporable getters in this paper. The microfluidic chip features microchannels with a height of around 100 nm. It is encapsulated with standard tube fittings, with leakage of less than 1 × 10−13 Pa ∙ m3∙ s−1. The conductance of the flowmeter is 10−12 m3∙ s−1, and the upper-pressure limit of the molecular flow is 105 Pa. It can control the intake flow of the adapted constant pressure test system from 10−11 to 10−7 Pa ∙ m3∙ s−1. Using this system, we tested the hydrogen adsorption capability of the Zr–Fe getter at different working pressures/temperatures and the types of gas it adsorbs were analysed. The results showed that the adsorbent has a noticeable adsorption effect on H2 and a partial adsorption effect on H2O, CO and CO2. SignificanceThe microfluidic chip flowmeter can provide a stable intake molecular flow for the adapted constant pressure test system. It ensures the reliability of the measurement results. The ability of the flowmeter to offer tiny flow rates at 105 Pa can drastically simplify the test system and is more user-friendly for getters tests with poor adsorption performance. It has positive significance for industrial research on the non-evaporable getter.

Rapid prototyping for high-pressure microfluidics

Soft lithography has permitted rapid prototyping of precise microfluidic features by patterning a deformable elastomer such as polydimethylsiloxane (PDMS) with a photolithographically patterned mold. In microfluidics applications where the flexibility of PDMS is a drawback, a variety of more rigid materials have been proposed. Compared to alternatives, devices fabricated from epoxy and glass have superior mechanical performance, feature resolution, and solvent compatibility. Here we provide a detailed step-by-step method for fabricating rigid microfluidic devices from soft lithography patterned epoxy and glass. The bonding protocol was optimized yielding devices that withstand pressures exceeding 500 psi. Using this method, we demonstrate the use of rigid high aspect ratio spiral microchannels for high throughput cell focusing.

Rapid microfluidics prototyping through variotherm desktop injection molding for multiplex diagnostics.

In this work, we demonstrate an inexpensive method of prototyping microfluidics using a desktop injection molding machine. A centrifugal microfluidic device with a novel central filling mechanism was developed to demonstrate the technique. We overcame the limitations of desktop machines in replicating microfluidic features by variotherm heating and cooling the mold between 50 °C and 110 °C within two minutes. Variotherm heating enabled good replication of microfeatures, with a coefficient of variation averaging only 3.6% attained for the measured widths of 100 μm wide molded channels. Using this methodology, we produced functional polystyrene centrifugal microfluidic chips, capable of aliquoting fluids into 5.0 μL reaction chambers with 97.5% accuracy. We performed allele-specific loop-mediated isothermal amplification (AS-LAMP) reactions for genotyping CYP2C19 alleles on these chips. Readouts were generated using optical pH sensors integrated onto chips, by drop-casting sensor precursor solutions into reaction chambers before final chip assembly. Positive reactions could be discerned by decreases in pH sensor fluorescence, thresholded against negative control reactions lacking the primers for nucleic acid amplification and with time-to-results averaging 38 minutes. Variotherm desktop injection molding can enable researchers to prototype microfluidic devices more cost-effectively, in an iterative fashion, due to reduced costs of smaller, in-house molds. Designs prototyped this way can be directly translated to mass production, enhancing their commercialization potential and positive impacts.

Diode Laser and Polyimide Tape Enables Cheap and Fast Fabrication of Flexible Microfluidic Sensing Devices.

Wearable devices are a new class of healthcare monitoring devices designed for use in close contact with the patient's body. Such devices must be flexible to follow the contours of human anatomy. With numerous potential applications, a wide variety of flexible wearable devices have been created, taking various forms and functions. Therefore, different fabrication techniques and materials are employed, resulting in fragmentation of the list of equipment and materials needed to make different devices. This study attempted to simplify and streamline the fabrication process of all key components, including microfluidic chip and flexible electrode units. A combination of diode laser CNC machine and polyimide tape is used to fabricate flexible microfluidic chip and laser-induced graphene (LIG) electrodes, to create flexible microfluidic sensing devices. Laser ablation on polyimide tape can directly create microfluidic features on either PDMS substrates or LIG electrodes. The two components can be assembled to form a flexible microfluidic sensing device that can perform basic electrochemical analysis and conform to curved surfaces while undergoing microfluidic flow. This study has shown that simple, commonly available equipment and materials can be used to fabricate flexible microfluidic sensing devices quickly and easily, which is highly suitable for rapid prototyping of wearable devices.

Ring Cross-over Based GA for DFMB Chip Design and Medical Image Compression

: Currently, the clinical data stored in the cloud is easily accessible, and the patient’s data can be shared among treatment centers. In such a case, to handle additional data, the cloud data must be of a lesser scale. A process of compression was introduced to minimize the data with no losing data in order to achieve this size reduction. This paper conducts the experiment in two approaches: fast routing operations and compression from the chip in the DMFB approach. To apply this process of compression, the collected data from the chip was transformed into an image, and then compression of the image was performed utilizing a genetic algorithm (GA) based on a ring crossover. Consequently, the biochip of the 8x8 array is integrated into the power and area with the ring cross-module for an effective energy consumption operation. The technique of the process is utilized by the Microfluidic (MF) feature to handle and maintain the droplets. Also, the optimization process is performed by combining related pin actuation segments in parallel and the control pin to prevent pin-actuation conflicts. Through the optimization process, it synchronizes the length. This proposed approach decreases the consumption of the power and area. The outcome of the simulation indicates an increase in dynamic power, static power, and delay. Image compression is performed with the aid of this algorithm. In addition, for better outcomes, this GA compression application was contrasted with wavelet compressions.

Merits and advances of microfluidics in the pharmaceutical field: design technologies and future prospects

Microfluidics is used to manipulate fluid flow in micro-channels to fabricate drug delivery vesicles in a uniform tunable size. Thanks to their designs, microfluidic technology provides an alternative and versatile platform over traditional formulation methods of nanoparticles. Understanding the factors that affect the formulation of nanoparticles can guide the proper selection of microfluidic design and the operating parameters aiming at producing nanoparticles with reproducible properties. This review introduces the microfluidic systems’ continuous flow (single-phase) and segmented flow (multiphase) and their different mixing parameters and mechanisms. Furthermore, microfluidic approaches for efficient production of nanoparticles as surface modification, anti-fouling, and post-microfluidic treatment are summarized. The review sheds light on the used microfluidic systems and operation parameters applied to prepare and fine-tune nanoparticles like lipid, poly(lactic-co-glycolic acid) (PLGA)-based nanoparticles as well as cross-linked nanoparticles. The approaches for scale-up production using microfluidics for clinical or industrial use are also highlighted. Furthermore, the use of microfluidics in preparing novel micro/nanofluidic drug delivery systems is presented. In conclusion, the characteristic vital features of microfluidics offer the ability to develop precise and efficient drug delivery nanoparticles.

Derivation and Differentiation of Human Pluripotent Stem Cells in Microfluidic Devices.

An integrative approach based on microfluidic design and stem cell biology enables capture of the spatial-temporal environmental evolution underpinning epigenetic remodeling and the morphogenetic process. We examine the body of literature that encompasses microfluidic applications where human induced pluripotent stem cells are derived starting from human somatic cells and where human pluripotent stem cells are differentiated into different cell types. We focus on recent studies where the intrinsic features of microfluidics have been exploited to control the reprogramming and differentiation trajectory at the microscale, including the capability of manipulating the fluid velocity field, mass transport regime, and controllable composition within micro- to nanoliter volumes in space and time. We also discuss studies of emerging microfluidic technologies and applications. Finally, we critically discuss perspectives and challenges in the field and how these could be instrumental for bringing about significant biological advances in the field of stem cell engineering.

Capturing individual charged microparticles and cells with microelectrodes in a circular microfluidic channel

Capture and, subsequently, in situ characterization of microparticles generally require complex fabrication processes and sophisticated techniques in microfluidic devices. In this paper, we present a simple and inexpensive technique for capturing and applying chemical agents to individual charged microparticles with the same glass microelectrode in a circular microfluidic channel. The microfluidic device features a thin poly-dimethyl-siloxane (PDMS) layer (100 µm, thickness) above the circular channel (diameter, 50 µm) so that the tip of the glass microelectrode can enter into the channel by penetrating through the thin PDMS layer. When an electric field is applied across the tip of the microelectrodes, the tip can capture charged microparticles or living cells that are flowing through the channel. Meanwhile, the same glass microelectrode can be used to apply chemical agents to the captured particles or living cells by filling the solution of the chemical agent into its tip. This simple design method offered simple fabrication without the need for any specialized equipment for the manipulation of charged particles or living cells in controlled microenvironments.

Characterization of a Centrifugal Microfluidic Orthogonal Flow Platform.

To bring to bear the power of centrifugal microfluidics on vertical flow immunoassays, control of flow orthogonally through nanoporous membranes is essential. The on-disc approach described here leverages the rapid print-cut-laminate (PCL) disc fabrication and prototyping method to create a permanent seal between disc materials and embedded nanoporous membranes. Rotational forces drive fluid flow, replacing capillary action, and complex pneumatic pumping systems. Adjacent microfluidic features form a flow path that directs fluid orthogonally (vertically) through these embedded membranes during assay execution. This method for membrane incorporation circumvents the need for solvents (e.g., acetone) to create the membrane-disc bond and sidesteps issues related to undesirable bypass flow. In other recently published work, we described an orthogonal flow (OF) platform that exploited embedded membranes for automation of enzyme-linked immunosorbent assays (ELISAs). Here, we more fully characterize flow patterns and cellulosic membrane behavior within the centrifugal orthogonal flow (cOF) format. Specifically, high-speed videography studies demonstrate that sample volume, membrane pore size, and ionic composition of the sample matrix significantly impact membrane behavior, and consequently fluid drainage profiles, especially when cellulosic membranes are used. Finally, prototype discs are used to demonstrate proof-of-principle for sandwich-type antigen capture and immunodetection within the cOF system.

Cyclic swelling enabled, electrically conductive 3D porous structures for microfluidic urinalysis devices

Urinalysis is a simple and non-invasive approach for the diagnosis and monitoring of organ health and also is often used as a facile technique in assessment of substance abuse. However, quantitative urinalysis is predominantly limited to clinical laboratories. Here, we present an electrical sensing based, reusable, cellular microfluidic device that offers a fast urinalysis through quantitative reading of the electrical signals. The spatial soft porous scaffolds decorated with electrically conductive multiwalled carbon nanotubes that are capable of physically interacting with biomarkers in urine are developed through a cyclic swelling/absorption process of soft materials and are utilized to manufacture the cellular microfluidic device. The sensing capability, sensitivity and reusability (via sunlight exposure) of the device to monitor red blood cells, Escherichia coli, and albumin are systemically demonstrated by programming mechanical deformation of porous scaffolds. Ex vivo experiments in disease mouse models confirm the diagnosis robustness of the device in comparable results with existing biochemical tests. The full integration of electrically conductive nanomaterials into soft scaffolds provides a foundation for devising bioelectronic devices with mechanically programmable microfluidic features in a low-cost manner, with broad applications for rapid disease diagnoses through body fluid.

Differential Impedance Sensing platform for high selectivity antibody detection down to few counts: A case study on Dengue Virus

We developed a biosensing system for serological detection of viruses based on the impedance variation between gold microelectrodes upon the capture of the target antibodies hybridized with nanobeads for signal amplification. The microfluidic platform core features a Differential Impedance Sensing (DIS) architecture between a reference and an active sensor able to reach nanoparticle resolution of few tens. The biosensor, functionalized with a copoly layer housing a synthetic peptide probe, has shown a limit of detection (LOD) below 100 pg/mL using a model IgG antibody spiked in a buffer. The biosensor was also tested with human serum samples for quantitative counts of anti-Dengue Virus antibodies, reaching a sensitivity that outperforms commercial ELISA kit. The system is perfectly suited to be easily reconfigured for novel probes by simply modifying the preparation of the biosensor chip surface, thus addressing a wide range of pathogens and diseases with clinically relevant concentrations for rapid immunoassays in a point of care setting.

Multi-material digital light processing bioprinting of hydrogel-based microfluidic chips

Recent advancements in digital-light-processing (DLP)-based bioprinting and hydrogel engineering have enabled novel developments in organs-on-chips. In this work, we designed and developed a multi-material, DLP-based bioprinter for rapid, one-step prototyping of hydrogel-based microfluidic chips. A composite hydrogel bioink based on poly-ethylene-glycol-diacrylate (PEGDA) and gelatin methacryloyl (GelMA) was optimized through varying the bioprinting parameters such as light exposure time, bioink composition, and layer thickness. We showed a wide range of mechanical properties of the microfluidic chips for various ratios of PEGDA:GelMA. Microfluidic features of hydrogel-based chips were then tested using dynamic flow experiments. Human-derived tumor cells were encapsulated in 3D bioprinted structures to demonstrate their bioactivity and cell-friendly environment. Cell seeding experiments then validated the efficacy of the selected bioinks for vascularized micro-tissues. Our biofabrication approach offers a useful tool for the rapid integration of micro-tissue models into organs-on-chips and high-throughput drug screening platforms.

Spatially and optically tailored 3D printing for highly miniaturized and integrated microfluidics

Traditional 3D printing based on Digital Light Processing Stereolithography (DLP-SL) is unnecessarily limiting as applied to microfluidic device fabrication, especially for high-resolution features. This limitation is due primarily to inherent tradeoffs between layer thickness, exposure time, material strength, and optical penetration that can be impossible to satisfy for microfluidic features. We introduce a generalized 3D printing process that significantly expands the accessible spatially distributed optical dose parameter space to enable the fabrication of much higher resolution 3D components without increasing the resolution of the 3D printer. Here we demonstrate component miniaturization in conjunction with a high degree of integration, including 15 μm × 15 μm valves and a 2.2 mm × 1.1 mm 10-stage 2-fold serial diluter. These results illustrate our approach’s promise to enable highly functional and compact microfluidic devices for a wide variety of biomolecular applications.

Ultrasensitive Multiparameter Phenotyping of Rare Cells Using an Integrated Digital-Molecular-Counting Microfluidic Well Plate.

Integrated microfluidic cellular phenotyping platforms provide a promising means of studying a variety of inflammatory diseases mediated by cell-secreted cytokines. However, immunosensors integrated in previous microfluidic platforms lack the sensitivity to detect small signals in the cellular secretion of proinflammatory cytokines with high precision. This limitation prohibits researchers from studying cells secreting cytokines at low abundance or existing at a small population. Herein, the authors present an integrated platform named the "digital Phenoplate (dPP)," which integrates digital immunosensors into a microfluidic chip with on-chip cell assay chambers, and demonstrates ultrasensitive cellular cytokine secretory profile measurement. The integrated sensors yield a limit of detection as small as 0.25 pg mL-1 for mouse tumor necrosis factor alpha (TNF-α). Each on-chip cell assay chamber confines cells whose population ranges from ≈20 to 600 in arrayed single-cell trapping microwells. Together, these microfluidic features of the dPP simultaneously permit precise counting and image-based cytometry of individual cells while performing parallel measurements of TNF-α released from rare cells under multiple stimulant conditions for multiple samples. The dPP platform is broadly applicable to the characterization of cellular phenotypes demanding high precision and high throughput.