Continuous-flow Microfluidics Research Articles

Hybrid Digital-Droplet Microfluidic Chip for Applications in Droplet Digital Nucleic Acid Amplification: Design, Fabrication and Characterization

Microfluidic-based platforms have become a hallmark for chemical and biological assays, empowering micro- and nano-reaction vessels. The fusion of microfluidic technologies (digital microfluidics, continuous-flow microfluidics, and droplet microfluidics, just to name a few) presents great potential for overcoming the inherent limitations of each approach, while also elevating their respective strengths. This work exploits the combination of digital microfluidics (DMF) and droplet microfluidics (DrMF) on a single substrate, where DMF enables droplet mixing and further acts as a controlled liquid supplier for a high-throughput nano-liter droplet generator. Droplet generation is performed at a flow-focusing region, operating on dual pressure: negative pressure applied to the aqueous phase and positive pressure applied to the oil phase. We evaluate the droplets produced with our hybrid DMF–DrMF devices in terms of droplet volume, speed, and production frequency and further compare them with standalone DrMF devices. Both types of devices enable customizable droplet production (various volumes and circulation speeds), yet hybrid DMF–DrMF devices yield more controlled droplet production while achieving throughputs that are similar to standalone DrMF devices. These hybrid devices enable the production of up to four droplets per second, which reach a maximum circulation speed close to 1540 µm/s and volumes as low as 0.5 nL.

Recent development of microfluidics-based platforms for respiratory virus detection.

With the global outbreak of SARS-CoV-2, the inadequacies of current detection technology for respiratory viruses have been recognized. Rapid, portable, accurate, and sensitive assays are needed to expedite diagnosis and early intervention. Conventional methods for detection of respiratory viruses include cell culture-based assays, serological tests, nucleic acid detection (e.g., RT-PCR), and direct immunoassays. However, these traditional methods are often time-consuming, labor-intensive, and require laboratory facilities, which cannot meet the testing needs, especially during pandemics of respiratory diseases, such as COVID-19. Microfluidics-based techniques can overcome these demerits and provide simple, rapid, accurate, and cost-effective analysis of intact virus, viral antigen/antibody, and viral nucleic acids. This review aims to summarize the recent development of microfluidics-based techniques for detection of respiratory viruses. Recent advances in different types of microfluidic devices for respiratory virus diagnostics are highlighted, including paper-based microfluidics, continuous-flow microfluidics, and droplet-based microfluidics. Finally, the future development of microfluidic technologies for respiratory virus diagnostics is discussed.

Nucleation control of quantum dot synthesis in a microfluidic continuous flow reactor

The use of microfluidics in chemical synthesis is topical due to the potential to improve reproducibility and the ability promptly interrogate a wide range of reaction parameters, the latter of which is necessary for the training of artificial intelligence (AI) algorithms. Applying microfluidic techniques to semiconductor nanocrystals, or quantum dots (QDs), is challenging due to the need for a high-temperature nucleation event followed by particle growth at lower temperatures. Such a high-temperature gradient can be realized using complex, segmented microfluidic reactor designs, which represents an engineering approach. Here, an alternative chemical approach is demonstrated using the cluster seed method of nanoparticle synthesis in a simple microfluidic reactor system. This enables quantum dot nucleation at lower temperatures due to the presence of molecular organometallic compounds (NMe4)4[Cd10Se4(SPh)16] and (NMe4)4[Zn10Se4(SPh)16]. This integration of cluster seeding with microfluidics affords a new mechanism to tailor the reaction conditions for optimizing yields and tuning product properties.

Design Automation for Continuous-Flow Lab-on-a-Chip Systems: A One-Pass Paradigm

Owing to the high complexity of chip architecture and assay protocol, considerable effort has been directed toward the design automation of continuous-flow microfluidics over the past decade. Existing methods, however, perform the corresponding design tasks, including binding, scheduling, placement, and routing separately, leading to serious gaps between different steps and potentially even cause design failure. To overcome these drawbacks, in this article, we propose a one-pass design paradigm for continuous-flow microfluidic lab-on-a-chip systems, integrating all the design steps into an “organic whole,” which has never been considered in prior work. With the proposed paradigm, all the design tasks can be synchronized seamlessly and performed in a combined manner, thereby eliminating the gaps between design steps. Consequently, optimized biochip architectures can be generated without any design adjustments and modifications. The experimental results demonstrate the effectiveness of the proposed automation flows.

Continuous flow microfluidics for colloidal particle assembly on porous substrates.

Self-assembly of colloidal particles for 'bottom-up' fabrication of various patterns and structures is critical for a range of applications including, but not limited to, energy generation and storage, nanomaterial structures, biomimetics, and biosensing. Multiple self-assembly techniques, such as substrate templating-via topological or chemical patterning-and solvent evaporation were discussed in our previous papers and have been developed for the deposition of patterned self-assembled structures, such as bands of colloidal particles, on various substrates. While the templating techniques are limited in applications due to the requirements for pattern-specific prior substrate engineering to fabricate the desired structure, solvent evaporation requires longer assembly times and precise control over environmental conditions. In this paper, a template-free, continuous flow process, which is facilitated by continuous solvent drainage through porous substrates, is demonstrated for the self-assembly of colloidal particles into high-aspect ratio (>103, length to width) structures, such as linear arrays or grid structures. Colloidal particles were assembled both on polymeric and metallic porous membranes, with rapid assembly times.

Microfluidic solvent extraction of no-carrier-added 64Cu from irradiated Zn target for radiopharmaceutical preparation

Microfluidics based liquid-liquid extraction is emerging as an enabling and powerful technology in radiochemical separations of medically important radioisotopes. In this study, we report the production of high specific activity 64Cu via 64Zn (n, p) 64Cu reaction in a research reactor. A microfluidic continuous flow liquid-liquid extraction system based on the use of dithizone as extractant was developed for efficient separation of miniscule quantity of 64Cu (in ng level) from bulk irradiated Zn target (in g level). The extraction process is completely selective for 64Cu, which could subsequently be retrieved in the desired medium for preparation of radiopharmaceuticals. The overall yield of 64Cu after the separation process was > 85 % and it was obtained with adequately high radionuclidic, radiochemical and chemical purities. Additionally, the effective specific activity of radio chemically separated 64Cu was determined to be 12.2 ± 0.8 GBq/µg, which is quite close to that reported for cyclotron produced no-carrier-added (NCA) 64Cu available commercially. The final 64Cu formulation could be effectively used for preparation of a variety of radiopharmaceuticals with high radiolabeling yields. The promising results obtained in this study open the pathway to achieve continuous and efficient recovery of reactor produced NCA 64Cu from the irradiated target using an automated microfluidic setup.

MiniControl 2.0: Co-Synthesis of Flow and Control Layers for Microfluidic Biochips With Strictly Constrained Control Ports

Recent advances in continuous-flow microfluidics have enabled highly integrated lab-on-a-chip biochips. These chips can execute complex biochemical applications precisely and efficiently within a tiny area, but they require a large number of control ports and the corresponding control logic to generate required pressure patterns for flow control, which, consequently, offset their advantages and prevent their wide adoption. In this article, we propose the first flow-control layer co-synthesis flow called MiniControl, for continuous-flow microfluidic biochips under strict constraints for control ports, incorporating high-level synthesis, physical design, and control system design simultaneously, which has never been considered in previous work. With the maximum number of allowed control ports specified in advance, this synthesis flow aims to generate biochip architectures with high execution efficiency and the corresponding control systems with optimized timing performance. Besides, the overall cost of a biochip can be reduced and the tradeoff between a control system and execution efficiency of biochemical applications can be evaluated for the first time. The experimental results demonstrate that MiniControl leads to high execution efficiency, low platform cost, as well as excellent timing performance, while strictly satisfying the given control-port constraints.

Microfluidic Continuous Flow DNA Fragmentation based on a Vibrating Sharp-tip.

Fragmentation of DNA into short fragments is of great importance for detecting and studying DNAs. Current microfluidic methods of DNA fragmentation are either inefficient for generating small fragments or rely on microbubbles. Here, we report a DNA fragmentation method in a 3D-printed microfluidic device, which allows efficient continuous flow fragmentation of genomic DNAs without the need for microbubbles. This method is enabled by localized acoustic streaming induced by a single vibrating sharp-tip. Genomic DNAs were fragmented into 700 to 3000 bp fragments with a low power consumption of ~140 mW. The system demonstrated successful fragmentation under a wide range of flow rates from 1 to 50 μL/min without the need for air bubbles. Finally, the utility of the continuous DNA fragmentation method was demonstrated to accelerate the DNA hybridization process for biosensing. Due to the small footprint, continuous flow and bubble-free operation, and high fragmentation efficiency, this method demonstrated great potential for coupling with other functional microfluidic units to achieve an integrated DNA analysis platform.

A Lab-in-a-Fiber optofluidic device using droplet microfluidics and laser-induced fluorescence for virus detection

Microfluidics has emerged rapidly over the past 20 years and has been investigated for a variety of applications from life sciences to environmental monitoring. Although continuous-flow microfluidics is ubiquitous, segmented-flow or droplet microfluidics offers several attractive features. Droplets can be independently manipulated and analyzed with very high throughput. Typically, microfluidics is carried out within planar networks of microchannels, namely, microfluidic chips. We propose that fibers offer an interesting alternative format with key advantages for enhanced optical coupling. Herein, we demonstrate the generation of monodisperse droplets within a uniaxial optofluidic Lab-in-a-Fiber scheme. We combine droplet microfluidics with laser-induced fluorescence (LIF) detection achieved through the development of an optical side-coupling fiber, which we term a periscope fiber. This arrangement provides stable and compact alignment. Laser-induced fluorescence offers high sensitivity and low detection limits with a rapid response time making it an attractive detection method for in situ real-time measurements. We use the well-established fluorophore, fluorescein, to characterize the Lab-in-a-Fiber device and determine the generation of sim 0.9 nL droplets. We present characterization data of a range of fluorescein concentrations, establishing a limit of detection (LOD) of 10 nM fluorescein. Finally, we show that the device operates within a realistic and relevant fluorescence regime by detecting reverse-transcription loop-mediated isothermal amplification (RT-LAMP) products in the context of COVID-19 diagnostics. The device represents a step towards the development of a point-of-care droplet digital RT-LAMP platform.

Mixing during Trapping Enabled a Continuous-Flow Microfluidic Smartphone Immunoassay Using Acoustic Streaming.

Smartphone-enabled microfluidic chemiluminescence immunoassay is a promising portable system for point-of-care (POC) biosensing applications. However, due to the rather faint emitted light in such a limited sample volume, it is still difficult to reach the clinically accepted range when the smartphone serves as a standalone detector. Besides, the multiple separation and washing steps during sample preparation hinder the immunoassay's applications for POC usage. Herein, we proposed a novel acoustic streaming tweezers-enabled microfluidic immunoassay, where the probe particles' purification, reaction, and sensing were simply achieved on the same chip at continuous-flow conditions. The dedicatedly designed high-speed microscale vortexes not only enable dynamic trapping and washing of the probe particles on-demand but also enhance the capture efficiency of the heterogeneous particle-based immunoassay through active mixing during trapping. The enriched probe particles and enhanced biomarker capture capability increase the local chemiluminescent light intensity and enable direct capture of the immunobinding signal by a regular smartphone camera. The system was tested for prostate-specific antigen (PSA) sensing both in buffer and serum, where a limit of detection of 0.2 ng/mL and a large dynamic response range from 0.3 to 10 ng/mL using only 10 μL of sample were achieved in a total assay time of less than 15 min. With the advantages of on-chip integration of sample preparation and detection and high sensing performance, the developed POC platform could be applied for many on-site diagnosis applications.

Sensing of inorganic ions in microfluidic devices

The past two decades witnessed the widespread adoption of microfluidics for miniaturized and automated chemical analysis. Microfluidic techniques require less sample, consume less reagents, and may enhance the sensitivity, precision, response time, and throughput of the analytical methods. The integration of multiple components and functionalities in microfluidic chips can further lead to fully automated Micro Total Analysis Systems (Lab-on-a-Chip). This review focuses on chemical and biochemical sensing of inorganic ions including electrolyte cations, heavy metal ions, and small anions in microfluidic devices. We provide an overview on how different optical and electrochemical ion sensing schemes based on liquid sensing phases or solid sensing phases can be coupled with microfluidics. Various microfluidics platforms including continuous-flow microfluidics, droplet microfluidics, digital microfluidics, centrifugal microfluidics, cell-based microfluidics, paper-based microfluidics, and wearable microfluidics are covered. The sensing principle, chip material, device fabrication, detector configuration, and analytical performance are presented for selected systems. This review concludes with a brief outlook on how microfluidic ion sensors may be further advanced.

Eliminating air bubble in microfluidic systems utilizing integrated in-line sloped microstructures.

In most microfluidic systems, formation and accumulation of air and other gas bubbles can be detrimental to their operation. Air bubbles in a microfluidic channel induce a pressure profile fluctuation and therefore disturb the stability of the system. Once an air bubble is generated, itis also extremely difficult to remove such bubbles from the microfluidic systems. In tissue and cell culture microfluidic systems, a single air bubble can completely shear off cells that are being cultured. Air bubbles can be especially problematic in microfluidic systems that have to operate for long periods of time, since completely eliminating the generation of air bubbles for prolonged periods of time, where a single air bubble can ruin an entire multi-day/multi-week experiment, is extremely challenging. Several in-line and off-chip bubble traps have been developed so far, but cannot completely eliminate air bubbles from the system or are relatively difficult to integrate into microfluidic systems. Recent advancements in two-photon polymerization (2PP)-based microfabrication method eliminates the restriction in Z-axis control in conventional two-dimensional microfabricationmethods, and thus enables complex 3D structures to be fabricated at sub-micrometer resolution. In this work, by utilizing this 2PP technique, we developed a sloped microfluidic structure that is capable of both trapping and real-time removal of air bubbles from the system in a consistent and reliable manner. The novel structures and designs developed in this work present a unique opportunity to overcome many limitations of currentmethods, bringstate-of-the-art solutions in air bubble removal, and enable a multifunctional microfluidic device to operate seamlessly free from air bubble disruption. The microfabricated system was tested in both droplet microfluidics and continuous-flow microfluidics applications, and demonstrated to be effective in preventing air bubble aggregation over time. This simple sloped microstructure can be easily integrated into broad ranges ofmicrofluidic devices to minimize bubble introduction, which will contribute to creating a stable and bubble-free microfluidic platform amenable for long-term operation.

Design and Comparison of Resonant and Non-Resonant Single-Layer Microwave Heaters for Continuous Flow Microfluidics in Silicon-Glass Technology

This paper presents a novel concept for the co-design of microwave heaters and microfluidic channels for sub-microliter volumes in continuous flow microfluidics. Based on the novel co-design concept, two types of heaters are presented, co-designed and manufactured in high-resistivity silicon-glass technology, resulting in a building block for consumable and mass-producible micro total analysis systems. Resonant and non-resonant co-planar waveguide transmission line heaters are investigated for heating of sub-micro-liter liquid volumes in a channel section at 25 GHz. The heating rates of 16 and 24 °C/s are obtained with power levels of 32 dBm for the through line and the open-ended line microwave heater, respectively. The heating uniformity of developed devices is evaluated with a Rhodamine B and deionized water mixture on a micrometer scale using the microwave-optical measurement setup. Measurement results showed a good agreement with simulations and demonstrated the potential of microwave heating for microfluidics.

Operation and Optimization of Microwave-Heated Continuous-Flow Microfluidics

Microwave (MW) technology can be powerful for electrification and process intensification but limited fundamental understanding of scalability and design principles hinders its effective use. In th...

Metered reagent injection into microfluidic continuous flow sampling for conductimetric ocean dissolved inorganic carbon sensing

Continuous autonomous measurement of total dissolved inorganic carbon (TCO2) in the oceans is critical for climate change modelling and ocean acidification measurement. A microfluidic conductivity-based approach will permit integration of miniaturised chemical analysis systems into Argo ocean floats, for long-term, high-accuracy depth profiling of dissolved CO2 with minimal reagent payload. Precise metering, suitable for sample acidification and CO2 liberation, is addressed. Laser etched microfluidic snake channel restrictors and asymmetric Y-meters were fabricated, with channel dimensions down to ∼75 μm, to adjust metering ratios between seawater and acid simulants. Hydrodynamic resistances, from flow versus pressure measurements, were compared with finite element simulations for various cross-section profiles and areas. Microfluidic metering circuits were constructed from various resistance snake channels and Y-junction components. Sample to acid volume ratios (meter ratio) up to 100:1 have been achieved with 300 μm wide snake channels for lengths >m. At highest resolution, the footprint would be >600 mm2. Circuits based solely on asymmetric Y-junctions gave maximum meter ratios of 16:1 with a footprint of <40 mm2 and ∼0.2% precision. Further refinement is required to ensure the integrity of such small channels in integration of metering units into full TCO2 analysis microfluidic circuits.

Preparation of Peptides with High Affinity to Cancer Targets in MRNA Display via Continuous-Flow Microfluidics

Preparation of Peptides with High Affinity to Cancer Targets in MRNA Display via Continuous-Flow Microfluidics

Vortex Fluidic Ethenolysis, Integrating a Rapid Quench of Ruthenium Olefin Metathesis Catalysts

Ruthenium-catalysed ethenolysis occurs in a vortex fluidic device (VFD) – a scalable, thin-film microfluidic continuous flow process. This process takes advantage of the efficient mass transfer of gaseous reagents into the dynamic thin film of liquid. Also reported is the rapid quenching of the ruthenium-based olefin metathesis catalyst by the addition of a saturated solution of N-acetyl-l-cysteine in MeCN, as a convenient alternative to previously reported quenching methods.

Micro-nanoparticles magnetic trap: Toward high sensitivity and rapid microfluidic continuous flow enzyme immunoassay.

In this work, we developed a microfluidic system for immunoassays where we combined the use of magnetic nanoparticles as immunosupport, a microfluidic magnetic trap, and a fluorogenic substrate in continuous flow for detection which, together with the optimization of the functionalization of surfaces to minimize nonspecific interactions, resulted in a detection limit in the order of femtomolar and a total assay time of 40 min for antibiotin antibody detection. A magnetic trap made of carbonyl-iron microparticles packaged inside a 200 m square microchannel was used to immobilize and concentrate nanoparticles. We functionalized the surface of the iron microparticles with a silica-polyethylene glycol (PEG) shell to avoid corrosion and unspecific protein binding. A new one-step method was developed to coat acrylic microchannels with an organofunctional silane functionalized with PEG to minimize unspecific binding. A model immunoassay was performed using nanoparticles decorated with biotin to capture antibiotin rabbit Immunoglobulin G (IgG) as target primary antibody. The detection was made using antirabbit IgG labeled with the enzyme alkaline phosphatase as a secondary antibody, and we measured fluorescence with a fluorescence microscope. All steps of the immunoassay were performed inside the chip. A calibration curve was obtained in which a detection limit of 8 pg/ml of antibiotin antibody was quantified. The simplicity of the device and the fact that it is made of acrylic, which is compatible with mass production, make it ideal for Point-Of-Care applications.

Microfluidic continuous flow synthesis of 1,5-ditosyl-1,5-diazocane-3,7-dione using response surface methodology

3,3,7,7-tetrakis(difluoroamino) octahydro-1,5-dinitro-1,5-diazocine (HNFX) is a high-density energetic oxidizer with four difluoroamino groups (-NF2). In this study, 1,5-ditosyl-1,5-diazocane-3,7-dione, as an important intermediate for synthesis of HNFX, was successfully synthesized using Swern oxidation in a traditional batch reactor and a continuous flow microreactor, respectively. 1,5-dotosyl-1,5-diazocane-3,7-dione was characterized by HPLC, FTIR, 1H NMR, 13C NMR, Mass spectrometry, and X-ray crystal diffraction. Compared with the traditional batch reactor, the microreactor showed several advantages, including less reaction time, milder reaction temperature, higher yield and selectivity for 1,5-ditosyl-1,5-diazocane-3,7-dione. Moreover, the microreactor could ensure the safer and large-scale industrial production of 1,5-ditosyl-1,5-diazocane-3,7-dione. However, some solids produced in Swern oxidation which might block the channels (diameter of 0.3mm) in the microreactor. To overcome the challenges, the experimental device was modified to suit for Swern oxidation, contributing to wider application of the microreactor. Besides, response surface methodology (RSM) was introduced and an appropriate mathematical model was built to optimize experimental conditions. The optimum experimental parameters were recommended as 7.8℃ for the reaction temperature, 7.7mL/min for the flow rate, and 6% for the concentration of oxalyl chloride. The actual yield of 1,5-ditosyl-1,5-diazocane-3,7-dione was 89.7%, which was in great agreement with the highest predicted yield (90.1%).

The Application of Microfluidics in Preparing Nano Drug Delivery Systems

The Application of Microfluidics in Preparing Nano Drug Delivery Systems