Droplet Generation Technique Research Articles

PH-induced morphological reversible transition from microparticles to vesicles for effective bacteria entrapment

Polymer particles with stimuli-responsive properties offer promising applications in healthcare, chemical reactors, development of artificial cells and organelles, as well as in the entrapment of bacteria. In this study, a novel biocompatible, biodegradable, and pH-responsive diblock copolymer based on polylactide (PLA) and poly(2-(diisopropylamino)ethyl methacrylate) (PDPA) was synthesized via a metal-free one-pot/simultaneous ring-opening polymerization (ROP) and reversible addition-fragmentation chain transfer (RAFT) approach (ROP/RAFT). This copolymer was then employed to produce highly monodisperse microparticles using microfluidics droplet generation technique. Utilizing confocal microscopy imaging, a reversible pH-induced morphological transition from microparticles to vesicle-like structures was observed at pH 5.1. The reversible shift in morphology from microparticles to giant vesicles (Vs) in acidic environments is triggered by the protonation of amino groups of the PDPA block, rendering vesicle surfaces positively charged − an advantageous feature for attracting and engulfing negatively charged bacteria. Initial validation involved electrostatic interactions with negatively charged latex resin beads followed by assessing interaction capabilities with gram-negative bacteria, Escherichia coli (E. coli). Additionally, the reversible morphological transition of microparticles-to-vesicles was employed to study drug release at different pHs. This approach proven to be a promising strategy for targeted drug delivery and bacteria entrapment using smart pH-responsive microparticles.

A Magnetically Driven Biodegradable Microsphere with Mass Production Capability for Subunit Vaccine Delivery and Enhanced Immunotherapy.

Subunit vaccines have emerged as a promising strategy in immunotherapy for combating viral infections and cancer. Nevertheless, the clinical application of subunit vaccines is hindered by limitations in antigen delivery efficiency, characterized by rapid clearance and inadequate cellular uptake. Here, a novel subunit vaccine delivery system utilizing ovalbumin@magnetic nanoparticles (OVA@MNPs) encapsulated within biodegradable gelatin methacryloyl (GelMA) microspheres was proposed to enhance the efficacy of antigen delivery. OVA@MNPs-loaded GelMA microspheres, denoted as OMGMs, can be navigated through magnetic fields to deliver subunit vaccines into the lymphatic system efficiently. Moreover, the biodegradable OMGMs enabled the sustained release of subunit vaccines, concentrating OVA around lymph nodes and enhancing the efficacy of induced immune response. OMGMs were produced through a microfluidic droplet generation technique, enabling mass production. In murine models, OMGMs successfully accumulated antigens in lymph nodes abundant in antigen-presenting cells, leading to enhanced cellular and humoral immunity and pronounced antitumor effects with a single booster immunization. In conclusion, these findings highlight the promise of OMGMs as a practical subunit vaccination approach, thus addressing the limitations associated with antigen delivery efficiency and paving the way for advanced immunotherapeutic strategies.

All-In-One OsciDrop Digital PCR System for Automated and Highly Multiplexed Molecular Diagnostics.

Digital PCR (dPCR) holds immense potential for precisely detecting nucleic acid markers essential for personalized medicine. However, its broader application is hindered by high consumable costs, complex procedures, and restricted multiplexing capabilities. To address these challenges, an all-in-one dPCR system is introduced that eliminates the need for microfabricated chips, offering fully automated operations and enhanced multiplexing capabilities. Using this innovative oscillation-induced droplet generation technique, OsciDrop, this system supports a comprehensive dPCR workflow, including precise liquid handling, pipette-based droplet printing, in situ thermocycling, multicolor fluorescence imaging, and machine learning-driven analysis. The system's reliability is demonstrated by quantifying reference materials and evaluating HER2 copy number variation in breast cancer. Its multiplexing capability is showcased with a quadruplex dPCR assay that detects key EGFR mutations, including 19Del, L858R, and T790M in lung cancer. Moreover, the digital stepwise melting analysis (dSMA) technique is introduced, enabling high-multiplex profiling of seven major EGFR variants spanning 35 subtypes. This innovative dPCR system presents a cost-effective and versatile alternative, overcoming existing limitations and paving the way for transformative advances in precision diagnostics.

Microfluidic Study of Oil Droplet Stability in Produced Water with Combinations of Production Chemicals

Discharged produced water contains traces of hydrocarbons after separation that might affect the eco-toxicity. These hydrocarbons can be oil droplets that are stabilized by added production chemicals and their combinations. The chemicals highly affect the treatment of the produced water before either reinjection or discharge to the sea. We apply microfluidic droplet generation for rapid evaluation of the synergistic effects of the combinations of production chemicals on the oil droplet stability. The coalescence frequency of dispersed droplets is calculated on the basis of high-speed camera video acquisition. The dispersed phase is either a model oil or a bottomhole crude oil sample, both in an aqueous medium. The individual and combined effects of a commercial demulsifier and a corrosion inhibitor are evaluated on the basis of the hydrophilic–lipophilic deviation theory. The presence of as low as 4 ppm of the water-in-oil demulsifier is demonstrated to stabilize oil-in-water droplets, while the corrosion inhibitor creates droplets in oil–water slug flow fashion. The presence of the corrosion inhibitor in conjunction with the demulsifier results in the droplet adherence to the channel surface. The importance of the right dosage and the right combination of production chemicals is demonstrated. The microfluidic droplet generation technique is a valuable tool for assessing the effects of production chemicals on oil droplet stability.

Picoliter Droplet Generation and Dense Bead-in-Droplet Encapsulation via Microfluidic Devices Fabricated via 3D Printed Molds.

Picoliter-scale droplets have many applications in chemistry and biology, such as biomolecule synthesis, drug discovery, nucleic acid quantification, and single cell analysis. However, due to the complicated processes used to fabricate microfluidic channels, most picoliter (pL) droplet generation methods are limited to research in laboratories with cleanroom facilities and complex instrumentation. The purpose of this work is to investigate a method that uses 3D printing to fabricate microfluidic devices that can generate droplets with sizes <100 pL and encapsulate single dense beads mechanistically. Our device generated monodisperse droplets as small as ~48 pL and we demonstrated the usefulness of this droplet generation technique in biomolecule analysis by detecting Lactobacillus acidophillus 16s rRNA via digital loop-mediated isothermal amplification (dLAMP). We also designed a mixer that can be integrated into a syringe to overcome dense bead sedimentation and found that the bead-in-droplet (BiD) emulsions created from our device had <2% of the droplets populated with more than 1 bead. This study will enable researchers to create devices that generate pL-scale droplets and encapsulate dense beads with inexpensive and simple instrumentation (3D printer and syringe pump). The rapid prototyping and integration ability of this module with other components or processes can accelerate the development of point-of-care microfluidic devices that use droplet-bead emulsions to analyze biological or chemical samples with high throughput and precision.

Fabrication of Nanowalled Catalytically Self-Threaded Supramolecular Polyrotaxane Microcapsules Using Droplet Microfluidics

Micrometer-scale monodisperse droplets are produced to generate nanowalled supramolecular microcapsules using microfluidics for high reproducibility and high-throughput manipulation, efficient material consumption, and control over hierarchical structure, shape, and size. In this study, an optimized microfluidic droplet generation technique and a unique liquid–liquid interfacial polymerization method were applied to fabricate the monodisperse polyrotaxane–based supramolecular microcapsules in a fast and simple way. To minimize the uncertainty due to droplet volume variation, the inlet pressures were supplied from the same source while lowering the interfacial tension and the main channel hydrodynamic resistance, which are critical for high monodispersity. The target polyrotaxane network (PN) was simply formed at the interface of the water and oil phases in ultra-monodisperse microdroplets via the cucurbit[6]uril (CB6)-catalyzed azide–alkyne cycloaddition (CB6-AAC) reaction between azido- and alkyne-functionalized tetraphenylporphyrin monomers (TPP-4AZ and TPP-4AL). The thickness of the interfacially assembled PN microcapsules was 20 nm as analyzed by cross-sectional TEM and TEM-EDX techniques. The resultant water-in-oil PN microcapsules were highly monodisperse in size and able to retain target molecules. Here, rhodamine 6G (Rh6G)-loaded PN microcapsules were fabricated, and the release rate of the Rh6G cargo was investigated over time for controlled drug release applications.

3차원 물방울 조각 생성장치의 구현을 위한 물방울 생성기법

사단법인 한국컴퓨터그래픽스학회의 논문지로서 그래픽스 분야와 그에 연관된 연구영역의 새로운 발견과 최첨단 연구결과를 발표하는 토론의 장입니다. 학회를 대표하는 학술지인 논문지는 1995년에 논문지 발간을 시작으로 2000년부터 3월, 6월, 9월, 12월 총 4회 발간되었고, 2015년부터는 KCGS 학술대회 특별호를 7월에 발간하면서 총 5회 발간되고 있습니다.

High-throughput single-cell whole-genome amplification through centrifugal emulsification and eMDA

Single-cell whole-genome sequencing (scWGS) is mainly used to probe intercellular genomic variations, focusing on the copy number variations or alterations and the single-nucleotide variations (SNVs) occurring within single cells. Single-cell whole-genome amplification (scWGA) needs to be applied before scWGS but is challenging due to the low copy number of DNA. Besides, many genomic variations are rare within a population of cells, so the throughput of currently available scWGA methods is far from satisfactory. Here, we integrate a one-step micro-capillary array (MiCA)-based centrifugal droplet generation technique with emulsion multiple displacement amplification (eMDA) and demonstrate a high-throughput scWGA method, MiCA-eMDA. MiCA-eMDA increases the single-run throughput of scWGA to a few dozen, and enables the assessment of copy number variations and alterations at 50-kb resolution. Downstream target enrichment further enables the detection of SNVs with 20% allele drop-out.

A digital acoustofluidic device for on-demand and oil-free droplet generation

We report a digital acoustofluidic device for on-demand and oil-free droplet generation. By applying a programmed radio frequency signal to a circular interdigital transducer, the dynamic focused acoustic pressure profiles generated rise up and dispense sample liquids from a reservoir to dynamically eject the droplets into the air. Our device allows droplets to be dispensed on demand with precisely controlled generation time and sequence, and accurate droplet volume. Moreover, we also demonstrate the generation of a droplet with a volume of 24 pL within 10 ms, as well as the encapsulation of a single cell into droplets. This acoustofluidic droplet generation technique is simple, biocompatible, and enables the on-demand droplet generation and encapsulation of many different biological materials with precise control, which is promising for single cell sampling and analysis applications.

Oscillating dispersed-phase co-flow microfluidic droplet generation: Multi-droplet size effect.

Controllable generation of microdroplets at desired sizes and throughputs is important in many applications. Many biological assays require size-optimized droplets for effective encapsulation of analytes and reagents. To perform size optimization, different-size droplets must be generated from identical sources of samples to prevent potential cross-sample variations or other sources of error. In this paper, we introduce a novel alteration of the co-flow droplet generation technique to achieve multi-size generation of monodispersed droplets. Using a custom-made mechanism, we oscillate the disperse-phase (d-phase) flow nozzle perpendicular to the continuous phase (c-phase) flow in a co-flow channel. Oscillation of the d-phase nozzle introduces an additional lateral drag force to the growing droplets while exposing them to various levels of axial drag owing to the parabolic velocity distribution of the c-phase flow. Superimposing both effects results in simultaneous and repeatable generation of monodispersed droplets with different sizes. The effect of nozzle oscillation frequency (f = 0-15 Hz) on droplet generation at different d-phase (Qd = 0.05, 0.10, and 0.50 ml/min) and c-phase (Qc = 2, 5, and 10 ml/min) flow rates was studied. A wide range of monodispersed droplets (4nl-4 μl) were generated using this method. Droplet sizes were directly proportional to the We number and inversely proportional to the Ca number and oscillation frequency. Our technique is promising for applications such as aqueous two-phase systems, where due to inherently low interfacial tension, the d-phase flow forms a long stable jet which can be broken into droplets using the additional oscillatory drag in our device.

Freezing on a Chip—A New Approach to Determine Heterogeneous Ice Nucleation of Micrometer-Sized Water Droplets

We are presenting a new approach to analyze the freezing behavior of aqueous droplets containing ice nucleating particles. The freezing chip consists of an etched and sputtered (15 × 15 × 1) mm gold-plated silicon or pure gold chip, enabling the formation of droplets with defined diameters between 20 and 80 µm. Several applications like an automated process control and an automated image evaluation were implemented to improve the quality of heterogeneous freezing experiments. To show the functionality of the setup, we compared freezing temperatures of aqueous droplets containing ice nucleating particles (i.e., microcline, birch pollen washing water, juniper pollen, and Snomax® solution) measured with our setup, with literature data. The ice nucleation active surface/mass site density (ns/m) of microcline, juniper pollen, and birch pollen washing water are shown to be in good agreement with literature data. Minor variations can be explained by slight differences in composition and droplet generation technique. The nm values of Snomax® differ by up to one order of magnitude at higher subzero temperatures when compared with fresh samples but are in agreement when compared with reported data of aged Snomax® samples.

Oscillating dispersed-phase co-flow microfluidic droplet generation: jet length reduction effect

Microdroplet generation methods are assessed by two important criteria of droplet throughput and size dispersity. The widely-used co-flow droplet generation technique is bottlenecked with droplet polydispersity at high throughputs due to transition to an unstable jetting regime at high dispersed-phase (d-phase) flow rates. In this paper, we introduce a novel technique to oscillate the d-phase nozzle inside the continuous phase (c-phase) channel to suppress the jetting effect. The effect of the nozzle oscillation frequency (0-15 Hz) on the jet length was studied at different d-phase (Qd = 1.8, 2.4 and 3.0 ml min-1) and c-phase (Qc = 6, 12 and 18 ml min-1) flow rates and d-phase viscosities (1, 2.5, and 6 mPa s). The jet length was directly proportional to the d-phase flow rate and inversely proportional to the oscillation frequency. Oscillation-induced jet length reduction was more significant at high jet velocities, but a less steep jet length reduction was always observed at oscillation frequencies higher than 10 Hz. A maximum jet length reduction of 70.8% was obtained at the highest d-phase and lowest c-phase flow rates. Increasing the viscosity of the d-phase resulted in diminishing the effect of oscillation on jet length reduction. Moreover, we observed that nozzle oscillation could disintegrate the long jet into droplets of various sizes that were mostly smaller than the stationary-mode droplets. We hypothesize that oscillating the dispersion nozzle at lower flow rates, without the jetting effect, can simultaneously generate multi-size monodisperse droplets. This active technique can also be implemented into aqueous two-phase systems (ATPSs) in which droplet generation is a difficult task.

Heparin-conjugated alginate multilayered microspheres for controlled release of bFGF

In order to effectively immobilize and control release of basic fibroblast growth factor (bFGF) from alginate microspheres, heparin-conjugated alginate (H-Alg) was first synthesized by covalent binding. Then multilayered H-Alg microspheres (multilayered microspheres) were fabricated via an electrostatic droplet generation technique followed by a layer-by-layer (LbL) self-assembly technique. Several techniques such as Fourier transform infrared spectroscopy (FTIR), 1H-NMR, zeta potential analysis and scanning electron microscopy (SEM) were used to characterize the properties of H-Alg (FTIR and 1H-NMR) and multilayered microspheres (FTIR, zeta potential analysis and SEM). bFGF binding efficiency, release profiles of bFGF from multilayered microspheres and the biological activity of released bFGF were well investigated. It was found that the bFGF binding efficiency of H-Alg microspheres was increased up to five times higher than that of the alginate microspheres. Additionally, the release profiles of bFGF from multilayered microspheres were sustained for two weeks with relieved initial burst release, and the release rate to bFGF could be regulated by controlling the number of deposited layers. Importantly, the released bFGF still retained its biological activity as assessed by the in vitro proliferation of NIH-3T3 mouse fibroblasts. In conclusion, this study presented an easy yet effective method for the controlled, sustained release of heparin-binding growth factors, using polyelectrolyte multilayer-coated heparin-conjugated alginate microspheres.

Incorporation of lipid nanoparticles into calcium alginate beads and characterization of the encapsulated particles by differential scanning calorimetry

The aim of this study was to encapsulate lipid nanoparticle dispersions into calcium alginate microbeads and to investigate the feasibility of this incorporation as well as the stability of the enclosed nanoparticles. Two methods were used to prepare the lipid-containing microbeads by external gelation, i.e. an electrostatic droplet generation technique and a spraying method using the two-fluid spray nozzle of a spray dryer. The particle sizes of the hydrogel beads were determined with laser diffraction. The first method produced particles with median sizes between 330 and 1350 μm, depending on the needle applied. Using the second method smaller particles (∼45 μm) could be prepared. Small tyloxapol-stabilized trimyristin nanoparticles (<100 nm) could easily be incorporated into the differently sized alginate beads. The small size of the lipid nanoparticles leads to a typical melting behavior characterized by the occurrence of multiple discrete melting peaks in their differential scanning calorimetry (DSC) thermograms which enabled the characterization of the lipid particles within the microbeads. Thus, any negative influence of the preparation procedure on the lipid dispersions could be detected and it was possible to verify optimization options of the incorporation procedure. Finally, handling and shelf life of the hydrogel beads were improved by a freezing process.

Preparation of N, O-carboxymethyl chitosan-Human-Like Collagen Microspheres for Drug Delivery

In this investigation, the N, O-carboxymethyl chitosan (NOCC) was synthesized. A novel pH-sensitive hydrogel system composed of a water-soluble chitosan derivative (N,O-carboxymethyl chitosan, NOCC)and Human-like collagen (HLC) blended with phosphate was developed for controlling drug delivery and NOCC–HLC microspheres were prepared by the technology of electrostatic droplet generation technique. The release profiles of a model protein drug (bovine serum albumin, BSA) from test hydrogels were studied in simulated gastric and intestinal media.

Boundary element analysis of the droplet dynamics induced by spark-generated bubble

Abstract Spark bubble droplet generation (SBDG) method is the most recently developed drop-on-demand droplet generation technique where the oscillation of a spark-generated bubble near a circular aperture causes a single droplet quite smaller than the aperture to form and break off. This paper investigates the fluid dynamics of the droplet generated through a flat plate aperture as well as through the nozzle of an axisymmetric chamber using boundary element method and high-speed photography. The results revealed that the bottom wall of the chamber would strongly influence the behavior of the bubble oscillating inside. In addition, it was found that with the same normalized nozzle size and bubble distance from air–liquid interface, a relatively smaller and faster droplet is generated from the chamber nozzle as compared with that formed through the flat plate aperture. Furthermore, although the droplet size decreases, its pinch-off time was found to increase by increasing the size of chamber feeder canal.

Preparation and characterization of PEM-coated alginate microgels for controlled release of protein

In this study, calcium-alginate microgels coated with a polyelectrolyte multilayer (PEM) were fabricated as a controlled-release system. This system was constructed via an electrostatic droplet generation technique followed by a layer-by-layer (LbL) self-assembly technique. The electrostatic droplet generation technique was reported as an easy method of preparing microgels, due to their mild preparation conditions and ability to preserve the biological activity of the encapsulated drugs. With the LbL self-assembly technique, the PEM could be fabricated on the microgels attributed to the electrostatic attraction between positive-charged chitosan (Chi) and negative-charged dextran sulfate (Dex). The properties of the prepared microgels were investigated using dynamic laser scattering (DLS), scanning electron microscopy (SEM), x-ray photoelectron spectroscopy (XPS), Fourier transform infrared (FTIR) spectrum and zeta potential analyzer. In vitro release study indicated that the initial burst release of the bovine serum albumin (BSA) from PEM-coated microgels was less compared to the uncoated microgels (19% versus 31% in 24 h). In addition, the sustained release of BSA from the PEM-coated microgels was recorded up to 1 month without any damage to BSA integrity. Thus, our results demonstrated that the PEM-coated microgels not only prolonged the release time, but also relieved the initial burst problem to some degree and preserved the biological activity of the encapsulated drugs. Moreover, the release rate of BSA could be regulated by controlling the number of deposited layers. In conclusion, this study presented an easy yet effective method for the controlled, sustained release of biological macromolecules.

A droplet-based building block approach for bladder smooth muscle cell (SMC) proliferation

Tissue engineering based on building blocks is an emerging method to fabricate 3D tissue constructs. This method requires depositing and assembling building blocks (cell-laden microgels) at high throughput. The current technologies (e.g., molding and photolithography) to fabricate microgels have throughput challenges and provide limited control over building block properties (e.g., cell density). The cell-encapsulating droplet generation technique has potential to address these challenges. In this study, we monitored individual building blocks for viability, proliferation and cell density. The results showed that (i) SMCs can be encapsulated in collagen droplets with high viability (>94.2 ± 3.2%) for four cases of initial number of cells per building block (i.e. 7 ± 2, 16 ± 2, 26 ± 3 and 37 ± 3 cells/building block). (ii) Encapsulated SMCs can proliferate in building blocks at rates that are consistent (1.49 ± 0.29) across all four cases, compared to that of the controls. (iii) By assembling these building blocks, we created an SMC patch (5 mm × 5 mm × 20 µm), which was cultured for 51 days forming a 3D tissue-like construct. The histology of the cultured patch was compared to that of a native rat bladder. These results indicate the potential of creating 3D tissue models at high throughput in vitro using building blocks.

06/01250 Microgravity experiments on flame spread along fuel-droplet arrays using a new droplet-generation technique: Mikami, M. et al. Combustion and Flame, 2005, 141, (3), 241–252

06/01250 Microgravity experiments on flame spread along fuel-droplet arrays using a new droplet-generation technique: Mikami, M. et al. Combustion and Flame, 2005, 141, (3), 241–252

Microgravity experiments on flame spread along fuel-droplet arrays using a new droplet-generation technique

A new droplet-array generation technique achieved high quality and high reliability in microgravity experiments on multiple-droplet combustion. Each fuel droplet formed at the intersection of fine, X-shaped SiC fibers when liquid fuel was supplied through a fine glass tube. We aligned several sets of these X-shaped fibers and their corresponding fine glass tubes to form a droplet array. All the droplets in the array were simultaneously generated in a short time. In flame-spread experiments, a hot-wire igniter ignited an end droplet to initiate the flame spread along the array. We demonstrated microgravity experiments of droplet array combustion using the new droplet-array generation technique at a drop-experiment facility, MGLAB, in Japan. We successfully generated large droplets, which often fell off the fiber intersection in normal gravity, by using this method in microgravity. This technique is also effective in droplet-array combustion experiments using high-volatility fuel, where prevaporization is substantial. We compared the flame-spread rate and the flame-spread limit of these linear droplet arrays with results of an existing experiment, and discussed the effects of the suspending fiber on the flame spread.