Multi-step synthesis of nanoparticles performed on millisecond time scale in a microfluidic droplet-based system

This paper reports a plug-based, microfluidic method for performing multi-step chemical reactions with millisecond time-control. It builds upon a previously reported method where aqueous reagents were injected into a flow of immiscible fluid (fluorocarbons)(H. Song et al., Angew. Chem. Int. Ed., 2003, 42, 768). The aqueous reagents formed plugs--droplets surrounded and transported by the immiscible fluid. Winding channels rapidly mixed the reagents in droplets. This paper shows that further stages of the reaction could be initiated by flowing additional reagent streams directly into the droplets of initial reaction mixture. The conditions necessary for an aqueous stream to merge with aqueous droplets were characterized. The Capillary number could be used to predict the behavior of the two-phase flow at the merging junction. By transporting solid reaction products in droplets, the products were kept from aggregating on the walls of the microchannels. To demonstrate the utility of this microfluidic method it was used to synthesize colloidal CdS and CdS/CdSe core-shell nanoparticles.     

Millisecond kinetics on a microfluidic chip using nanoliters of reagents

This paper describes a microfluidic chip for performing kinetic measurements with better than millisecond resolution. Rapid kinetic measurements in microfluidic systems are complicated by two problems: mixing is slow and dispersion is large. These problems also complicate biochemical assays performed in microfluidic chips. We have recently shown (Song, H.; Tice, J. D.; Ismagilov, R. F. Angew. Chem., Int. Ed. 2003, 42, 768-772) how multiphase fluid flow in microchannels can be used to address both problems by transporting the reagents inside aqueous droplets (plugs) surrounded by an immiscible fluid. Here, this droplet-based microfluidic system was used to extract kinetic parameters of an enzymatic reaction. Rapid single-turnover kinetics of ribonuclease A (RNase A) was measured with better than millisecond resolution using sub-microliter volumes of solutions. To obtain the single-turnover rate constant (k = 1100 +/- 250 s(-1)), four new features for this microfluidics platform were demonstrated: (i) rapid on-chip dilution, (ii) multiple time range access, (iii) biocompatibility with RNase A, and (iv) explicit treatment of mixing for improving time resolution of the system. These features are discussed using kinetics of RNase A. From fluorescent images integrated for 2-4 s, each kinetic profile can be obtained using less than 150 nL of solutions of reagents because this system relies on chaotic advection inside moving droplets rather than on turbulence to achieve rapid mixing. Fabrication of these devices in PDMS is straightforward and no specialized equipment, except for a standard microscope with a CCD camera, is needed to run the experiments. This microfluidic platform could serve as an inexpensive and economical complement to stopped-flow methods for a broad range of time-resolved experiments and assays in chemistry and biochemistry.     

High-temperature microfluidic synthesis of CdSe nanocrystals in nanoliter droplets

The high-temperature synthesis of CdSe nanocrystals in nanoliter-volume droplets flowing in a perfluorinated carrier fluid through a microfabricated reactor is presented. A flow-focusing nanojet structure with a step increase in channel height reproducibly generated octadecene droplets in Fomblin Y 06/6 perfluorinated polyether at capillary numbers up to 0.81 and with a droplet:carrier fluid viscosity ratio of 0.035. Cadmium and selenium precursors flowing in octadecene droplets through a high-temperature (240-300 degrees C) glass microreactor produced high-quality CdSe nanocrystals, as verified by optical spectroscopy and transmission electron microscopy. Isolating the reaction solution in droplets prevented particle deposition and hydrodynamic dispersion, allowing the reproducible synthesis of nanocrystals at three different temperatures and four different residence times in the span of 4 h. Our synthesis of a wide range of nanocrystals at high temperatures, high capillary numbers, and low viscosity ratio illustrates the general utility of droplet-based microfluidic reactors to encapsulate nanoliter volumes of organic or aqueous solutions and to precisely control chemical or biochemical reactions.     

Surface modification of droplet polymeric microfluidic devices for the stable and continuous generation of aqueous droplets

Droplet microfluidics performed in poly(methyl methacrylate) (PMMA) microfluidic devices resulted in significant wall wetting by water droplets formed in a liquid-liquid segmented flow when using a hydrophobic carrier fluid such as perfluorotripropylamine (FC-3283). This wall wetting led to water droplets with nonuniform sizes that were often trapped on the wall surfaces, leading to unstable and poorly controlled liquid-liquid segmented flow. To circumvent this problem, we developed a two-step procedure to hydrophobically modify the surfaces of PMMA and other thermoplastic materials commonly used to make microfluidic devices. The surface-modification route involved the introduction of hydroxyl groups by oxygen plasma treatment of the polymer surface followed by a solution-phase reaction with heptadecafluoro-1,1,2,2-tetrahydrodecyl trichlorosilane dissolved in fluorocarbon solvent FC-3283. This procedure was found to be useful for the modification of PMMA and other thermoplastic surfaces, including polycyclic olefin copolymer (COC) and polycarbonate (PC). Angle-resolved X-ray photoelectron spectroscopy indicated that the fluorination of these polymers took place with high surface selectivity. This procedure was used to modify the surface of a PMMA droplet microfluidic device (DMFD) and was shown to be useful in reducing the wetting problem during the generation of aqueous droplets in a perfluorotripropylamine (FC-3283) carrier fluid and could generate stable segmented flows for hours of operation. In the case of PMMA DMFD, oxygen plasma treatment was carried out after the PMMA cover plate was thermally fusion bonded to the PMMA microfluidic chip. Because the appended chemistry to the channel wall created a hydrophobic surface, it will accommodate the use of other carrier fluids that are hydrophobic as well, such as hexadecane or mineral oils.     

Flow-focusing generation of monodisperse water droplets wrapped by ionic liquid on microfluidic chips: from plug to sphere

Generating droplets via microfluidic chips is a promising technology in microanalysis and microsynthesis. To realize room-temperature ionic liquid (IL)-water two-phase studies in microscale, a water-immiscible IL was employed as the continuous phase for the first time to wrap water droplets (either plugs or spheres) on flow-focusing microfluidic chips. The IL, 1-butyl-3-methylimidazolium hexafluorophosphate ([BMIM][PF6]), could wet both hydrophilic and hydrophobic channel surfaces because of its dual role of hydrophilicity/hydrophobicity and extremely high viscosity, thus offering the possibility of wrapping water droplets in totally hydrophilic (THI), moderately hydrophilic (MHI), and hydrophobic (HO) channels. The droplet shape could be tuned from plug to sphere, with the volume from 6.3 nL to 65 pL, by adding an orifice in the focusing region, rendering the hydrophilic channel surface hydrophobic, and suppressing the Uw/UIL ratio below 1.0. Three different breakup processes were defined and clarified, in which the sub-steady breakup and steady breakup were essential for the formation of plugs and spheric droplets, respectively. The influences of channel hydrophilicity/hydrophobicity on droplet formation were carefully studied by evaluating the wetting abilities of water and IL on different surfaces. The superiority of IL over water in wetting hydrophobic surface led to the tendency of forming small, spheric aqueous droplets in the hydrophobic channel. This IL-favored droplet-based system represented a high efficiency in water/IL extraction, in which rhodamine 6G was extracted from aqueous droplets to [BMIM][PF6] in the hydrophobic orifice-included (HO-OI) channel in 0.51 s.     

Microgram-scale testing of reaction conditions in solution using nanoliter plugs in microfluidics with detection by MALDI-MS

This paper describes a microfluidic system to screen and optimize organic reaction conditions on a submicrogram scale. The system uses discrete droplets (plugs) as microreactors separated and transported by a continuous phase of a fluorinated carrier fluid. Previously, we demonstrated the use of a microfabricated PDMS plug-based microfluidic system to perform assays and crystallization experiments in aqueous solutions with optical detection. Here, we developed an approach that does not require microfabrication of microfluidic devices, is applicable to synthetic reactions in organic solvents, and uses detection by MALDI-MS. As a demonstration, conditions for selective deacetylation of ouabain hexaacetate were tested, and the optimum conditions for mono-, bis-, or trisdeacetylation have been identified. These conditions were validated by scale-up reactions and isolating these potentially neurotoxic products. Mono- and bisdeacetylated products are unstable intermediates in the deacetylation and were isolated for the first time. This system enables no-loss handling of submicroliter volumes containing a few micrograms of a compound of interest. It could become valuable for investigating or optimizing reactions of precious substrates (e.g., products of long synthetic sequences and natural products that can be isolated only in small quantities).     

Biopolymer microparticle and nanoparticle formation within a microfluidic device

This paper reports a novel microfluidic method for the production of cross-linked alginate microparticles and nanoparticles. We describe a continuous process relying on both thermodynamic and hydrodynamic factors to form microdroplets. A rapid cross-linking reaction thereafter allows solidification of the polymer droplets either within the microfluidic device or "off-chip" to form alginate micro- and nanoparticles. Monodisperse droplets are generated by extruding an aqueous alginate solution using an axisymmetric flow-focusing design. As they flow downstream in the channel, due to water and the continuous phase being partially miscible, the water diffuses very slowly out of the polymeric droplets into the transport fluid, which causes the shrinkage of the drops and the condensation of the polymer phase. The resulting size of the solid particles depends on the polymer concentration and the ensuing balance between the kinetics of the cross-linking reaction and the volume loss due to solvent diffusion. This work details both a single-step microfluidic technique for the formation of alginate microparticles of sizes ranging from 1 to 50 microm via near-equilibrium solvent diffusion within a microfluidic device and thereafter a two-step method, which was shown to generate biopolymer nanoparticles of sizes ranging from 10 to 300 nm. These novel methodologies are extremely flexible and can be extended to the preparation of micro- and nanoparticles from a wide range of single or mixed synthetic and biologically derived polymers.     

Composite poly(dimethylsiloxane)/glass microfluidic system with an immobilized enzymatic particle-bed reactor and sequential sample injection for chemiluminescence determinations

A three-layer poly(dimethylsiloxane) (PDMS)/glass microfluidic system for performing on-chip solid-phase enzymatic reaction and chemiluminescence (CL) reaction was used for the determination of glucose as a model analyte. A novel method for the immobilization of controlled-pore-glass based reactive particles on PDMS microreactor beds was developed, producing an on-chip solid-phase reactor that featured large reactive surface and low flow impedance. Efficient mixing of reagent/sample/carrier streams was achieved by incorporating chaotic mixer structures in the microfluidic channels. A conventional sequential injection (SI) system was adapted for direct coupling with the microfluidic system, and combined with hydrostatic delivery of reagents to achieve efficient and reproducible sample introduction at 10 μl levels. A detection limit of 10 μM glucose (3σ), and a precision of 3.1% RSD (n=7, 0.2 mM glucose) were obtained using the SI-microfluidic-CL system integrated with a glucose oxidase (GOD) reactor. Carryover was <5% at a throughput of 20 samples/h.     

Preparation of Poly(vinyl alcohol) Microspheres Based on Droplet Microfluidic Technology

A new method for preparing poly (vinyl alcohol) (PVA) microspheres was developed by using droplet microfluidic technology. In the microfluidic chip, a large number of uniform, monodispersed PVA droplets were prepared quickly and continuously by using droplet formation technology, and the droplet preparation speed reached 7 per second. The size of the PVA droplets could be controlled by changing the injection flow rate of the two-phase fluid and the width of microfluidic channel. Then the PVA microspheres were formed by physical crosslinking. This method has high preparation efficiency and good monodispersity of the obtained microspheres. Moreover, the process does not require the incorporation of chemical crosslinking agents, avoiding interference with the inclusion material, and is well suited for applications such as drug carrier.     

Controlled production of monodisperse double emulsions by two-step droplet breakup in microfluidic devices

A microfluidic device having both hydrophobic and hydrophilic components is exploited for production of multiple-phase emulsions. For producing water-in-oil-in-water (W/O/W) dispersions, aqueous droplets ruptured at the upstream hydrophobic junction are enclosed within organic droplets formed at the downstream hydrophilic junction. Droplets produced at each junction could have narrow size distributions with coefficients of variation in diameter of less than 3%. Control of the flow conditions produces variations in internal/external droplet sizes and in the internal droplet number. Both W/O/W emulsions (with two types of internal droplets) and oil-in-water-in-oil emulsions were prepared by varying geometry and wettability in microchannels.     

Formation of droplets and bubbles in a microfluidic T-junction-scaling and mechanism of break-up

This article describes the process of formation of droplets and bubbles in microfluidic T-junction geometries. At low capillary numbers break-up is not dominated by shear stresses: experimental results support the assertion that the dominant contribution to the dynamics of break-up arises from the pressure drop across the emerging droplet or bubble. This pressure drop results from the high resistance to flow of the continuous (carrier) fluid in the thin films that separate the droplet from the walls of the microchannel when the droplet fills almost the entire cross-section of the channel. A simple scaling relation, based on this assertion, predicts the size of droplets and bubbles produced in the T-junctions over a range of rates of flow of the two immiscible phases, the viscosity of the continuous phase, the interfacial tension, and the geometrical dimensions of the device.     

Investigating the effects of precursor concentration and gelling parameters on droplet-based generation of Ca-Alginate microgels: identifying new stable modes of droplet formation

Droplet-based microfluidics is an attractive approach for producing microgels due to its high potential to control the size and shape of the particles and precisely entrap the substances within the hydrogel matrix. However, the microfluidic generation of monodisperse microgels with desired structures is still challenging. Indeed, the rheological and interfacial properties of the immiscible fluids, as well as the adopted gelling strategy, play important roles in microfluidic methods. Herein, sodium alginate droplets with different concentrations are generated via a microfluidic device with a flow-focusing unit. Besides, a combined in situ and ex situ strategy is optimized to crosslink sodium alginate droplets in the presence of calcium ions. The effects of alginate concentration and junction width in the flow focusing unit are investigated on droplet size and droplet formation regimes. It is observed that by increasing the alginate concentration, the dripping regime of droplet formation may be transformed to one of the binary dripping or quasijetting regimes. In the binary dripping regime, two successive different-sized droplets are generated in each period of droplet formation, which leads to low monodispersity in the collected droplets. However, the droplets produced in the quasijetting regime are interestingly monodisperse and also smaller than those of the dripping and binary dripping regimes. The breakup dynamics of the alginate thread is also analyzed with a computational fluid dynamics (CFD) code. This analysis discloses that the viscous stresses, as well as the viscous dissipation, have important roles in controlling the stable modes of droplet formation.     

Comparison of monodisperse droplet generation in flow-focusing devices with hydrophilic and hydrophobic surfaces

A thin flow-focusing microfluidic channel is evaluated for generating monodisperse liquid droplets. The microfluidic device is used in its native state, which is hydrophilic, or treated with OTS to make it hydrophobic. Having both hydrophilic and hydrophobic surfaces allows for creation of both oil-in-water and water-in-oil emulsions, facilitating a large parameter study of viscosity ratios (droplet fluid/continuous fluid) ranging from 0.05 to 96 and flow rate ratios (droplet fluid/continuous fluid) ranging from 0.01 to 2 in one geometry. The hydrophilic chip provides a partially-wetting surface (contact angle less than 90°) for the inner fluid. This surface, combined with the unusually thin channel height, promotes a flow regime where the inner fluid wets the top and bottom of the channel in the orifice and a stable jet is formed. Through confocal microscopy, this fluid stabilization is shown to be highly influenced by the contact angle of the liquids in the channel. Non-wetting jets undergo breakup and produce drops when the jet is comparable to or smaller than the channel thickness. In contrast, partially-wetting jets undergo breakup only when they are much smaller than the channel thickness. Drop sizes are found to scale with a modified capillary number based on the total flow rate regardless of wetting behavior.     

At-line bioprocess monitoring by immunoassay with rotationally controlled serial siphoning and integrated supercritical angle fluorescence optics

In this paper we report a centrifugal microfluidic “lab-on-a-disc” system for at-line monitoring of human immunoglobulin G (hIgG) in a typical bioprocess environment. The novelty of this device is the combination of a heterogeneous sandwich immunoassay on a serial siphon-enabled microfluidic disc with automated sequential reagent delivery and surface-confined supercritical angle fluorescence (SAF)-based detection. The device, which is compact, easy-to-use and inexpensive, enables rapid detection of hIgG from a bioprocess sample. This was achieved with, an injection moulded SAF lens that was functionalized with aminopropyltriethoxysilane (APTES) using plasma enhanced chemical vapour deposition (PECVD) for the immobilization of protein A, and a hybrid integration with a microfluidic disc substrate. Advanced flow control, including the time-sequenced release of on-board liquid reagents, was implemented by serial siphoning with ancillary capillary stops. The concentration of surfactant in each assay reagent was optimized to ensure proper functioning of the siphon-based flow control. The entire automated microfluidic assay process is completed in less than 30 min. The developed prototype system was used to accurately measure industrial bioprocess samples that contained 10 mg mL⁻¹ of hIgG.     

High-throughput rheology in a microfluidic device

High-throughput rheological measurements in a microfluidic device are demonstrated. A series of microrheology samples are generated as droplets in an immiscible spacer fluid using a microfluidic T-junction. The compositions of the sample droplets are continuously varied over a wide range. Rheology measurements are made in each droplet using multiple particle tracking microrheology. We review critical design and operating parameters, including the droplet size, flow rates and rapid fabrication methods. Validation experiments are performed by measuring the solution viscosity of glycerine and the biopolymer heparin as a function of concentration. Overall, the combination of microrheology with microfluidics maximizes the number of rheological measurements while simultaneously minimizing the sample preparation time and amount of material, and should be particularly suited to the characterization of scarce or expensive materials.     

Hierarchical Porous Polymer Beads Prepared by Polymerization-induced Phase Separation and Emulsion-template in a Microfluidic Device

Porous polymer beads（PPBs） containing hierarchical bimodal pore structure with gigapores and meso-macropores were prepared by polymerization-induced phase separation（PIPS） and emulsion-template technique in a glass capillary microfluidic device（GCMD）. Fabrication procedure involved the preparation of water-in-oil emulsion by emulsifying aqueous solution into the monomer solution that contains porogen. The emulsion was added into the GCMD to fabricate the（water-in-oil）-in-water double emulsion droplets. The flow rate of the carrier continuous phase strongly influenced the formation mechanism and size of droplets. Formation mechanism transformed from dripping to jetting and size of droplets decreased from 550 μm to 250 μm with the increase in flow rate of the carrier continuous phase. The prepared droplets were initiated for polymerization by on-line UV-irradiation to form PPBs. The meso-macropores in these beads were generated by PIPS because of the presence of porogen and gigapores obtained from the emulsion-template. The pore morphology and pore size distribution of the PPBs were investigated extensively by scanning electron microscopy and mercury intrusion porosimetry（MIP）. New pore morphology was formed at the edge of the beads different from traditional theory because of different osmolarities between the water phase of the emulsion and the carrier continuous phase. The morphology and proportion of bimodal pore structure can be tuned by changing the kind and amount of porogen.     

Microfluidic platform for on-demand generation of spatially indexed combinatorial droplets

We propose a highly versatile and programmable nanolitre droplet-based platform that accepts an unlimited number of sample plugs from a multi-well plate, performs digitization of these sample plugs into smaller daughter droplets and subsequent synchronization-free, robust injection of multiple reagents into the sample daughter droplets on-demand. This platform combines excellent control of valve-based microfluidics with the high-throughput capability of droplet microfluidics. We demonstrate the functioning of a proof-of-concept device which generates combinatorial mixture droplets from a linear array of sample plugs and four different reagents, using food dyes to mimic samples and reagents. Generation of a one dimensional array of the combinatorial mixture droplets on the device leads to automatic spatial indexing of these droplets, precluding the need to include a barcode in each droplet to identify its contents. We expect this platform to further expand the range of applications of droplet microfluidics to include applications requiring a high degree of multiplexing as well as high throughput analysis of multiple samples.     

Droplet-based microfluidics for dose–response assay of enzyme inhibitors by electrochemical method

A simple but robust droplet-based microfluidic system was developed for dose–response enzyme inhibition assay by combining concentration gradient generation method with electrochemical detection method. A slotted-vials array and a tapered tip capillary were used for reagents introduction and concentration gradient generation, and a polydimethylsiloxane (PDMS) microfluidic chip integrated with microelectrodes was used for droplet generation and electrochemical detection. Effects of oil flow rate and surfactant on electrochemical sensing were investigated. This system was validated by measuring dose–response curves of three types of acetylcholinesterase (AChE) inhibitors, including carbamate pesticide, organophosphorus pesticide, and therapeutic drugs regulating Alzheimer's disease. Carbaryl, chlorpyrifos, and tacrine were used as model analytes, respectively, and their IC₅₀ (half maximal inhibitory concentration) values were determined. A whole enzyme inhibition assay was completed in 6 min, and the total consumption of reagents was less than 5 μL. This microfluidic system is applicable to many biochemical reactions, such as drug screening and kinetic studies, as long as one of the reactants or products is electrochemically active.     

微流控芯片停流液-液萃取技术的研究

基于微流控芯片的液-液萃取技术的研究是目前微流控芯片分析领域内的重要研究方向之一,与传统液-液萃取系统相比,萃取系统微型化所带来的优势表现为显著降低试样与试剂的消耗(仅为传统系统的万分之一)、分析速度快、易实现操作自动化和分析系统集成化。目前,在已报道的基于微流