Processes for the 3D Printing of Hydrodynamic Flow-Focusing Devices.

Flow focusing is an important hydrodynamic technique for cytometric analysis, enabling the rapid study of cellular samples to identify a variety of biological processes. To date, the majority of flow-focusing devices are fabricated using conventional photolithography or flame processing of glass capillaries. This article presents a suite of low-cost, millifluidic, flow-focusing devices that were fabricated using a desktop sterolithgraphy (SLA) 3D printer. The suite of SLA printing strategies consists of a monolithic SLA method and a hybrid molding process. In the monolithic SLA approach, 1.3 mm square millifluidic channels were printed as a single piece. The printed device does not require any post processing, such as bonding or surface polishing for optical access. The hybrid molding approach consists of printing a mold using the SLA 3D printer. The mold is treated to an extended UV exposure and oven baked before using PDMS as the molding material for the channel. To demonstrate the viability of these channels, we performed a series of experiments using several flow-rate ratios to show the range of focusing widths that can be achieved in these devices. The experiments are validated using a numerical model developed in ANSYS.

Directed Assembly of Particles for Additive Manufacturing of Particle-Polymer Composites.

Particle-polymer dispersions are ubiquitous in additive manufacturing (AM), where they are used as inks to create composite materials with applications to wearable sensors, energy storage materials, and actuation elements. It has been observed that directional alignment of the particle phase in the polymer dispersion can imbue the resulting composite material with enhanced mechanical, electrical, thermal or optical properties. Thus, external field-driven particle alignment during the AM process is one approach to tailoring the properties of composites for end-use applications. This review article provides an overview of externally directed field mechanisms (e.g., electric, magnetic, and acoustic) that are used for particle alignment. Illustrative examples from the AM literature show how these mechanisms are used to create structured composites with unique properties that can only be achieved through alignment. This article closes with a discussion of how particle distribution (i.e., microstructure) affects mechanical properties. A fundamental description of particle phase transport in polymers could lead to the development of AM process control for particle-polymer composite fabrication. This would ultimately create opportunities to explore the fundamental impact that alignment has on particle-polymer composite properties, which opens up the possibility of tailoring these materials for specific applications.

3D printed imaging platform for portable cell counting.

Despite having widespread application in the biomedical sciences, flow cytometers have several limitations that prevent their application to point-of-care (POC) diagnostics in resource-limited environments. 3D printing provides a cost-effective approach to improve the accessibility of POC devices in resource-limited environments. Towards this goal, we introduce a 3D-printed imaging platform (3DPIP) capable of accurately counting particles and perform fluorescence microscopy. In our 3DPIP, captured microscopic images of particle flow are processed on a custom developed particle counter code to provide a particle count. This prototype uses a machine vision-based algorithm to identify particles from captured flow images and is flexible enough to allow for labeled and label-free particle counting. Additionally, the particle counter code returns particle coordinates with respect to time which can further be used to perform particle image velocimetry. These results can help estimate forces acting on particles, and identify and sort different types of cells/particles. We evaluated the performance of this prototype by counting 10 µm polystyrene particles diluted in deionized water at different concentrations and comparing the results with a commercial Beckman-Coulter Z2 particle counter. The 3DPIP can count particle concentrations down to ∼100 particles per mL with a standard deviation of ±20 particles, which is comparable to the results obtained on a commercial particle counter. Our platform produces accurate results at flow rates up to 9 mL h-1 for concentrations below 1000 particle per mL, while 5 mL h-1 produces accurate results above this concentration limit. Aside from performing flow-through experiments, our instrument is capable of performing static experiments that are comparable to a plate reader. In this configuration, our instrument is able to count between 10 and 250 cells per image, depending on the prepared concentration of bacteria samples (Citrobacter freundii; ATCC 8090). Overall, this platform represents a first step towards the development of an affordable fully 3D printable imaging flow cytometry instrument for use in resource-limited clinical environments.

Soft Ferrofluid Actuator Based on 3D-Printed Scaffold Removal.

Fabricating soft functional materials via additive manufacturing is an emerging field with a wide variety of applications due to their ability to respond to specifically engineered stimuli (e.g., mechanical, electrical, magnetic, chemical). This article describes an approach to engineering magnetically sensitive structures using three-dimensional printing of acrylonitrile butadiene styrene scaffolds. These scaffolds are encapsulated in polydimethylsiloxane (PDMS) and removed using organic solvents. The open channels that remain after removal are filled in with a ferrofluid to render the structure magnetically sensitive. A three-point flexural test shows that introducing a channel in this way only reduces the flexural modulus of the PDMS by a factor ∼8%. We perform magnetic deflection experiments on samples with three different channel diameters. Our results show a linear dependence between applied magnetic field strength and deflection. We also find that there is a minimum magnetic field strength that needs to be applied to achieve deflection. These results suggest that there is a minimum yield stress, beyond which deflection will occur. We perform experiments on a more complex channel geometry to find that there are multiple modes of deflection. A rational approach to channel design may enable us to tune the mechanical response and direct these actuators to undergo complex motion.

Acoustophoretic assembly of millimeter-scale Janus fibers.

This article presents a method for the assembly of millimeter-scale Janus fibers using acoustophoresis as an assembly mechanism. An acoustic flow cell mounted to a 3D printer combines acoustophoresis and additive manufacturing in a unique approach that allows for the assembly of textured Janus fibers. A dispersion consisting of polymethylmethacrylate (PMMA) filler particles in a UV curable polymer resin is passed through an acoustically excited capillary tube. To fundamentally understand this process, we develop a suspension balance model that accounts for acoustophoresis and concentration-driven shear-induced diffusion. Once assembled, the particle-polymer dispersion is cured using UV illumination to create a polymer composite fiber with particles immobilized on one side in a Janus-like configuration. The Janus fiber is observed to modify the light transmission profile when rotated on an optical microscope stage. Tensile measurements of the fiber show that the Young's modulus of the Janus fiber (50.5 MPa) is approximately twice that of a fiber fabricated from the polymer alone (24.7 MPa). The process we describe here could serve as a pathway for the fabrication of a variety of functional Janus fibers with possible applications to wearable textiles, soft robotics or surgical sutures.

Protocol for assembling micro- and nanoparticles in a viscous liquid above a vibrating plate.

In this protocol, we demonstrate the use of a vibrating plate to drive the assembly of micro- and nanoparticles as an approach to high-throughput, large-scale directed assembly in a viscous liquid. Vibration drives the assembly of glass bead microparticles and iron oxide nanoparticles in contact with water over an area of 6400 mm2. We use a scaling analysis to show that there is a competition between acoustic radiation force and vibration-generated fluid flow in a viscous medium, which determines particle transport characteristics. For assembly in a viscous liquid, we find close agreement between the observed experimental results when compared to a numerical solution of the 2D wave equation that describes plate displacement. This model indicates that microparticles migrate along displacement gradients towards displacement anti-nodes where the magnitude of displacement is maximum. We also observe that nanoparticles migrate toward displacement nodes where the magnitude of displacement is zero. •Cost-effective directed assembly technique without the need for microfabrication facilities•Large-scale assembly produces heterogeneously ordered structures on a vibrating substrate.

High-Throughput Acoustofluidic Self-Assembly of Colloidal Crystals.

Colloidal crystals are encountered in a variety of energy-harvesting applications, where they serve as waveguides or filters for electromagnetic and electro-optic energy. Techniques such as electric or magnetic assembly are used to assemble colloidal crystals, but are limited by crystal size, yield, and throughput. This article demonstrates the continuous, high-throughput assembly of two-dimensional (2D)-colloidal crystals in an acoustofluidic flow cell. The device is fabricated using off-the-shelf components and does not require a clean-room access. An experimental state diagram shows how the fluid flow rate and voltage applied to the piezoelectric element in our device can tune the crystal microstructure. Highly ordered colloidal crystals are continuously assembled in less than a minute with a throughput yield of several hundred particles per minute using this device. The acoustically assembled ordered 2D crystals are immobilized using a UV-curable resin and extracted as ordered polymer-particle fibers, demonstrating the ability of using acoustic fields to assemble ordered structures embedded in bulk materials. Particle tracking is used to construct the cross-channel particle distribution to understand the effect of acoustic compression on colloidal crystal assembly. Microparticle image velocimetry data is compared to a theoretical transport model to quantify the effect fluid flow and acoustic trapping has on the colloidal crystal ensemble.

Line-Focused Optical Excitation of Parallel Acoustic Focused Sample Streams for High Volumetric and Analytical Rate Flow Cytometry.

Flow cytometry provides highly sensitive multiparameter analysis of cells and particles but has been largely limited to the use of a single focused sample stream. This limits the analytical rate to ∼50K particles/s and the volumetric rate to ∼250 µL/min. Despite the analytical prowess of flow cytometry, there are applications where these rates are insufficient, such as rare cell analysis in high cellular backgrounds (e.g., circulating tumor cells and fetal cells in maternal blood), detection of cells/particles in large dilute samples (e.g., water quality, urine analysis), or high-throughput screening applications. Here we report a highly parallel acoustic flow cytometer that uses an acoustic standing wave to focus particles into 16 parallel analysis points across a 2.3 mm wide optical flow cell. A line-focused laser and wide-field collection optics are used to excite and collect the fluorescence emission of these parallel streams onto a high-speed camera for analysis. With this instrument format and fluorescent microsphere standards, we obtain analysis rates of 100K/s and flow rates of 10 mL/min, while maintaining optical performance comparable to that of a commercial flow cytometer. The results with our initial prototype instrument demonstrate that the integration of key parallelizable components, including the line-focused laser, particle focusing using multinode acoustic standing waves, and a spatially arrayed detector, can increase analytical and volumetric throughputs by orders of magnitude in a compact, simple, and cost-effective platform. Such instruments will be of great value to applications in need of high-throughput yet sensitive flow cytometry analysis.

Characterizing gelatin hydrogel viscoelasticity with diffusing colloidal probe microscopy.

In this study, we investigate viscoelasticity in gelatin hydrogels using diffusing colloidal probe microscopy (DCPM) to directly measure the elastic potential energy interaction between colloidal probes and the underlying viscoelastic media. Gelatin samples are prepared in four different concentrations between 0.3wt% and 0.6wt% to examine changes in viscoelasticity with concentration. A force balance describing the interaction between the colloidal probes and the hydrogel as a spring-damper system lead to a simple model for mean square displacement. A histogram of locations sampled by the colloidal probes is directly related to the elastic potential energy and the effective spring constant of the gelatin hydrogels. The effective spring constant is a fixed parameter used in the mean square displacement model to find effective viscosity. These parameters are comparable to viscoelastic parameters obtain by a microrheology analysis of two-dimensional mean square displacements. These results can serve as a guide for assessing hydrogel systems where viscoelastic properties are an important factor in biomaterial design.

Reconfigurable assemblies of Janus rods in AC electric fields.

We investigate the electric field-induced assembly of Janus colloids composed of silica rods patterned with gold patches in both side- and tip-coated motifs. These shape and chemically anisotropic particles assemble into reconfigurable chains, whose structure depends on patch location, AC electric field strength, and frequency.

Janus colloidal matchsticks.

We fabricated chemically and shape-anisotropic colloids composed of silica rods coated with gold tips using a multistep process involving electric-field alignment and crystallization, microcontact printing, and selective metallization. Through direct observation, we found that these "Janus matchsticks" self-assemble into multipods (bi-, tri-, and tetrapods) of varying coordination number and patch angle in aqueous solution.

Multiple electrokinetic actuators for feedback control of colloidal crystal size.

We report a feedback control method to precisely target the number of colloidal particles in quasi-2D ensembles and their subsequent assembly into crystals in a quadrupole electrode. Our approach relies on tracking the number of particles within a quadrupole electrode, which is used in a real-time feedback control algorithm to dynamically actuate competing electrokinetic transport mechanisms. Particles are removed from the quadrupole using DC-field mediated electrophoretic-electroosmotic transport, while high-frequency AC-field mediated dielectrophoretic transport is used to concentrate and assemble colloidal crystals. Our results show successful control of the size of crystals containing 20 to 250 colloidal particles with less than 10% error. Assembled crystals are characterized by their radius of gyration, crystallinity, and number of edge particles, and demonstrate the expected size-dependent properties. Our findings demonstrate successful ensemble feedback control of the assembly of different sized colloidal crystals using multiple actuators, which has broad implications for control over nano- and micro- scale assembly processes involving colloidal components.

kT-scale colloidal interactions in high frequency inhomogeneous AC electric fields. I. Single particles.

We report nonintrusive optical microscopy measurements of single micrometer-sized silica and polystyrene colloids in inhomogeneous AC electric fields as a function of field amplitude and frequency. By using a Boltzmann inversion of the time-averaged sampling of single particles within inhomogeneous electric fields, we sensitively measure induced dipole-field interactions on the kT energy scale and fN force scale. Measurements are reported for frequencies when the particle polarizability is greater and less than the medium, as well as the crossover between these conditions when dipole-field interactions vanish. For all cases, the measured interactions are well-described by theoretical potentials by fitting a nondimensional induced dipole-field magnitude. While silica dipole-field magnitudes are well-described by existing electrokinetic models, the polystyrene results suggest an anomalously high surface conductance. Sensitive measurements of dipole-field interactions in this work provide a basis to understand dipole-dipole interactions in particle ensembles in the same measurement geometry in part II.

kT-scale colloidal interactions in high-frequency inhomogeneous AC electric fields. II. Concentrated ensembles.

We report nonintrusive optical microscopy measurements of ensembles of polystyrene colloids in inhomogeneous AC electric fields as a function of field frequency and particle size. By using an inverse Monte Carlo (MC) simulation analysis of time-averaged particle microstructures, we sensitively measure induced dipole-dipole interactions on the kT energy scale. Measurements are reported for frequencies when the particle polarizability is greater and less than the medium, as well as the crossover between these conditions when dipole-dipole interactions vanish. By using measured single dipole-field interactions and associated parameters from Part I as input in the inverse analysis, the dipole-dipole interactions in this work are accurately modeled with no adjustable parameters for conditions away from the crossover frequency (i.e., |f(CM)| > 0). As dipolar interactions vanish at the crossover, a single frequency-dependent parameter is introduced to account for microstructures that appear to result from weak AC electro-osmotic flow induced interactions. By connecting quantitative measures of equilibrium microstructures and kT-scale dipole-field and dipole-dipole interactions, our findings provide a basis to understand colloidal assembly in inhomogeneous AC electric fields.

Interactions and microstructures in electric field mediated colloidal assembly.

We report video microscopy measurements and computer simulations of quasi-two-dimensional configurations of micron sized colloids in 1 MHz ac electric fields between coplanar thin film electrodes. Interactions of induced dipoles (IDs) with each other and inhomogeneous electric fields (IFs) as a function of concentration and field amplitude produced microstructures including confined hard disk fluids, oriented dipolar chains, and oriented hexagonal close packed crystals. Equilibrium measurements and analyses of single colloids within electric fields were used to directly measure ID-IF interactions in the absence of many body effects. Measurements of concentrated systems were characterized in terms of density profiles across the electrode gap and angular pair distribution functions. In concentrated measurements, an inverse Monte Carlo analysis was used to extract the ID-ID interaction. A single adjustable parameter consistently modified the ID-IF potential and the ID-ID potential to account for weakening of ID as the result of the local particle concentration and configuration.