Single Polystyrene Beads Research Articles

Exceeding 80% Efficiency of Single-Bead Encapsulation in Microdroplets through Hydrogel Coating-Assisted Close-Packed Ordering.

High-efficiency encapsulation of single microbeads in microdroplets is essential for droplet-based high-throughput analysis such as single-cell genomics and digital immunoassays. However, the demand has been hindered by the Poisson statistics of beads arbitrarily distributed in the droplet partitions. Although techniques such as inertial ordering have been proven useful to improve bead-loading efficiency, a general method that requires no advanced microfluidics and owns compatibility with diverse bead types is still highly desired. In this paper, we present hydrogel coating-assisted close-packed ordering, a simple strategy that improves the bead-loading efficiency to over 80%. In the strategy, the raw beads are coated with a thin layer of hydrogel to become slightly compressible and lubricious, so that they can be close-packed in a microfluidic device and loaded into droplets in a synchronized manner. We first show that the thin hydrogel coating can be realized conveniently through jetting microfluidics or vortex emulsification. When loading single 30-μm polystyrene beads, we experimentally determine an overall efficiency of 81% with the proposed hydrogel coating strategy. Of note, the strategy is not sensitive to the selection of raw beads and can tolerate their polydispersity. Using the strategy, we achieve a cell capture rate of 68.8% when co-encapsulating HEK293T cells and polydispersed barcoded beads for single-cell transcriptomics. Further sequencing results verify that the reversible hydrogel coating does not affect the RNA capture behavior of the encapsulated barcoded beads. Given its convenience and broad compatibility, we anticipate that our strategy can be applied to various droplet-based high-throughput assays to improve their efficiency drastically.

Controlled Transport of Individual Microparticles Using Dielectrophoresis.

A dielectrophoretic device employing a planar array of microelectrodes is designed for controlled transport of individual microparticles. By exciting the electrodes in sequence, a moving dielectrophoretic force is created that can drag a particle across the electrodes in a straight line. The electrode shapes are designed to counter any lateral drift of the trapped particle during transport. This facilitates single particle transport by creating a narrow two-dimensional corridor for the moving dielectrophoretic force to operate on. The design and analysis processes are discussed in detail. Numerical simulations are performed to calculate the electromagnetic field distribution and the generated dielectrophoretic force near the electrodes. The Langevin equation is used for analyzing the trajectory of a microparticle under the influence of the external forces. The simulations show how the designed electrode geometry produces the necessary lateral confinement required for successful particle transport. Finally, experimental results are presented showing controlled bidirectional linear transport of single polystyrene beads of radius 10 and 5 μm for a distances 840 and 1100 μm, respectively. The capabilities of the proposed platform make it suitable for micro total analysis systems (μTAS) and lab-on-a-chip (LOC) applications.

Continuous spatial self-cleaning in GRIN multimode fiber for self-referenced multiplex CARS imaging.

We demonstrate how spatial beam self-cleaning and supercontinuum generation in graded-index multimode optical fibers can be directly applied in multiplex coherent anti-Stokes Raman Scattering (M-CARS) spectroscopy. Although supercontinuum generation causes pump depletion mainly in the center of the beam, the partial recovery of the pump brightness due to self-cleaning may enable self-referenced M-CARS, with no additional delay lines to synchronize pump and Stokes waves. As a proof-of-principle, we report examples of imaging of single chemical compounds and polystyrene beads. The new scheme paves the way towards simpler M-CARS systems based on multimode fiber sources.

Deciphering single- and multi-particle trapping dynamics under femtosecond pulsed excitation with simultaneous spatial and temporal resolution

Recent theoretical and experimental studies have shed light on how laser trapping dynamics under femtosecond pulsed excitation are fine-tuned by optical and thermal nonlinearities. Here, we present experimental results of trapping of single and multiple polystyrene beads (of 1 μm diameter). We show how integration and synchronization of bright-field video microscopy with confocal detection of backscatter provide both spatial and temporal resolution required to capture intricate details of nonlinear trapping dynamics. Such spatiotemporal detection is promising to have far-reaching applications in exploring controlled laser trapping and manipulations harnessed by optical and thermal nonlinearities.

Individual Microparticle Manipulation Using Combined Electroosmosis and Dielectrophoresis through a Si3N4 Film with a Single Micropore.

Porous dielectric membranes that perform insulator-based dielectrophoresis or electroosmotic pumping are commonly used in microchip technologies. However, there are few fundamental studies on the electrokinetic flow patterns of single microparticles around a single micropore in a thin dielectric film. Such a study would provide fundamental insights into the electrokinetic phenomena around a micropore, with practical applications regarding the manipulation of single cells and microparticles by focused electric fields. We have fabricated a device around a silicon nitride film with a single micropore (2–4 µm in diameter) which has the ability to locally focus electric fields on the micropore. Single microscale polystyrene beads were used to study the electrokinetic flow patterns. A mathematical model was developed to support the experimental study and evaluate the electric field distribution, fluid motion, and bead trajectories. Good agreement was found between the mathematic model and the experimental data. We show that the combination of electroosmotic flow and dielectrophoretic force induced by direct current through a single micropore can be used to trap, agglomerate, and repel microparticles around a single micropore without an external pump. The scale of our system is practically relevant for the manipulation of single mammalian cells, and we anticipate that our single-micropore approach will be directly employable in applications ranging from fundamental single cell analyses to high-precision single cell electroporation or cell fusion.

Optical manipulation of a dielectric particle along polygonal closed-loop geometries within a single water droplet

We report a new method to optically manipulate a single dielectric particle along closed-loop polygonal trajectories by crossing a suite of all-fiber Bessel-like beams within a single water droplet. Exploiting optical radiation pressure, this method demonstrates the circulation of a single polystyrene bead in both a triangular and a rectangle geometry enabling the trapped particle to undergo multiple circulations successfully. The crossing of the Bessel-like beams creates polygonal corners where the trapped particles successfully make abrupt turns with acute angles, which is a novel capability in microfluidics. This offers an optofluidic paradigm for particle transport overcoming turbulences in conventional microfluidic chips.

High-spatial-resolution deep tissue imaging with spectral-domain optical coherence microscopy in the 1700-nm spectral band.

.We present three-dimensional (3-D) high-resolution spectral-domain optical coherence microscopy (SD-OCM) by using a supercontinuum (SC) fiber laser source with 300-nm spectral bandwidth (full-width at half-maximum) in the 1700-nm spectral band. By using low-coherence interferometry with SC light and a confocal detection scheme, we realized lateral and axial resolutions of 3.4 and in tissue (), respectively. This is, to the best of our knowledge, the highest 3-D spatial resolution reported among those of Fourier-domain optical coherence imaging techniques in the 1700-nm spectral band. In our SD-OCM, to enhance the imaging depth, a full-range method was implemented, which suppressed the formation of a coherent ghost image and allowed us to set the zero-delay position inside the samples. We demonstrated the 3-D high-resolution imaging capability of 1700-nm SD-OCM through the measurement of an interference signal from a mirror surface and imaging of a single 200-nm polystyrene bead and a pig thyroid gland. Deep tissue imaging at a depth of up to 1.8 mm was also demonstrated. This is the first demonstration of 3-D high-resolution SD-OCM in the 1700-nm spectral band.

Functionalization of Atomic Force Microscope Cantilevers with Single-T Cells or Single-Particle for Immunological Single-Cell Force Spectroscopy.

Atomic force microscopy based single cell force spectroscopy (AFM-SCFS) is a powerful tool for studying biophysical properties of living cells. This technique allows for probing interaction strengths and dynamics on a live cell membrane, including those between cells, receptor and ligands, and alongside many other variations. It also works as a mechanism to deliver a physical or biochemical stimulus on single cells in a spatiotemporally controlled manner, thus allowing specific cell activation and subsequent cellular events to be monitored in real-time when combined with live-cell fluorescence imaging. The key step in those AFM-SCFS measurements is AFM-cantilever functionalization, or in other words, attaching a subject of interest to the cantilever. Here, we present methods to modify AFM cantilevers with a single T cell and a single polystyrene bead respectively for immunological studies. The former involves a biocompatible glue that couples single T cells to the tip of a flat cantilever in a solution, while the latter relies on an epoxy glue for single bead adhesion in the air environment. Two immunological applications associated with each cantilever modification are provided as well. The methods described here can be easily adapted to different cell types and solid particles.

Mie scattering from strongly absorbing airborne particles in a photophoretic trap

High-precision light scattering measurements of single, absorbing aerosol particles pose a challenge as the strong attenuation of light results in featureless spectra. Using the Kramers-Kronig relations and broadband light scattering, we develop and implement a method for the retrieval of both the size and the complex refractive index of particles that are strongly absorbing across large spectral regions. This method is validated using both synthetic spectra and scattered light intensity measurements from single dye-doped polystyrene beads of known radii that are held using a photophoretic trap.

Raman imaging through multimode sapphire fiber.

We report on a sapphire fiber Raman imaging probe's use for challenging applications where access is severely restricted. Small-dimension Raman probes have been developed previously for various clinical applications because they show great capability for diagnosing disease states in bodily fluids, cells, and tissues. However, applications of these sub-millimeter diameter Raman probes were constrained by two factors: first, it is difficult to incorporate filters and focusing optics at such small scale; second, the weak Raman signal is often obscured by strong background noise from the fiber probe material, especially the most commonly used silica, which has a strong broad background noise in low wavenumbers (<500-1700 cm-1). Here, we demonstrate the thinnest-known imaging Raman probe with a 60 μm diameter Sapphire multimode fiber in which both excitation and signal collection pass through. This probe takes advantage of the low fluorescence and narrow Raman peaks of Sapphire, its inherent high temperature and corrosion resistance, and large numerical aperture (NA). Raman images of Polystyrene beads, carbon nanotubes, and CaSO4 agglomerations are obtained with a spatial resolution of 1 μm and a field of view of 30 μm. Our imaging results show that single polystyrene bead (~15 µm diameter) can be differentiated from a mixture with CaSO4 agglomerations, which has a close Raman shift.

Dipolar and quadrupolar mode dissipation of spherical probes spinning in vortex beam acoustical tweezers

In this work, the possibility of simultaneously trapping and rotating single polystyrene beads, or clusters, against gravity with an ultrasonic vortex beam is demonstrated. The induced rotation of a single particle is compared to a torque balance model accounting for the acoustic response of the particle. Two dominating dissipation mechanisms of the acoustic orbital angular momentum responsible for the observed rotation are examined. The first takes place in the bulk of the absorbing particle, while the second arises as the angular momentum flux is dissipated in the viscous boundary layer surrounding the particle. Importantly, even in the long-wavelength (Rayleigh) regime, it is crucial to fully model the dissipation of the dipolar and quadrupolar vibrational modes of the particle in a slightly viscous fluid such as water. Further results suggest that, while the induced outer (Eckart) rotational flow can be neglected in water, it can play an important role in other viscous and complex fluids.

Orbital Angular Momentum Transfer to Stably Trapped Elastic Particles in Acoustical Vortex Beams.

The controlled rotation of solid particles trapped in a liquid by an ultrasonic vortex beam is observed. Single polystyrene beads, or clusters, can be trapped against gravity while simultaneously rotated. The induced rotation of a single particle is compared to a torque balance model accounting for the acoustic response of the particle. The measured torque (∼10 pN m for a driving acoustic power ∼40 W/cm^{2}) suggests two dominating dissipation mechanisms of the acoustic orbital angular momentum responsible for the observed rotation. The first takes place in the bulk of the absorbing particle, while the second arises as dissipation in the viscous boundary layer in the surrounding fluid. Importantly, the dissipation processes affect both the dipolar and quadrupolar particle vibration modes suggesting that the restriction to the well-known Rayleigh scattering regime is invalid to model the total torque even for spheres much smaller than the sound wavelength. The findings show that a precise knowledge of the probe elastic absorption properties is crucial to perform rheological measurements with maneuverable trapped spheres in viscous liquids. Further results suggest that the external rotational steady flow must be included in the balance and can play an important role in other liquids.

Rapid structural analysis of nanomaterials in aqueous solutions

Rapid structural analysis of nanoscale matter in a liquid environment represents innovative technologies that reveal the identities and functions of biologically important molecules. However, there is currently no method with high spatio-temporal resolution that can scan individual particles in solutions to gain structural information. Here we report the development of a nanopore platform realizing quantitative structural analysis for suspended nanomaterials in solutions with a high z-axis and xy-plane spatial resolution of 35.8 ± 1.1 and 12 nm, respectively. We used a low thickness-to-diameter aspect ratio pore architecture for achieving cross sectional areas of analyte (i.e. tomograms). Combining this with multiphysics simulation methods to translate ionic current data into tomograms, we demonstrated rapid structural analysis of single polystyrene (Pst) beads and single dumbbell-like Pst beads in aqueous solutions.

Encapsulation of single cells into monodisperse droplets by fluorescence-activated droplet formation on a microfluidic chip

Random compartmentalization of cells by common droplet formation methods, i.e., T-junction and flow-focusing, results in low occupancy of droplets by single cells. To resolve this issue, a fluorescence-activated droplet formation method was developed for the on-command generation of droplets and encapsulation of single cells. In this method, droplets containing one cell were generated by switching on/off a two-phase hydrodynamic gating valve upon optical detection of single cells. To evaluate the developed method, flow visualization experiments were conducted with fluorescein. Results indicated that picoliter droplets of uniform sizes (RSD<4.9%) could be generated. Encapsulation of single fluorescent polystyrene beads demonstrated an average of 94.3% droplets contained one bead. Further application of the developed methods to the compartmentalization of individual HeLa cells indicated 82.5% occupancy of droplets by single cells, representing a 3 fold increase in comparison to random compartmentalization.

Whispering Gallery Microcavity Nanosensor with In-Line Interferometer

Here we demonstrate a simple but robust interferometry scheme that connects an ultra-high quality factor whispering gallery micro cavity to a FC/PC connectorized single mode fibre. With this scheme, single Polystyrene beads with a radius as small as 25 nm can be detected at a signal-to-noise ratio of around 3.

Creation of single-particle environment by positive dielectrophoresis and liquid dielectrophoresis

Two electrical mechanisms for manipulating particles and fluids, dielectrophoresis (DEP) and liquid dielectrophoresis (LDEP), are integrated in a microfluidic chip for creating the single-particle environment. The fluid is activated by LDEP with a 100-kHz/240-Vpp signal. When the single polystyrene bead approaches the trapping area, positive DEP force is utilized to capture and immobilize the bead. After trapping the bead, the process of liquid cutting and droplet creation is employed to create a droplet containing a single bead by LDEP with a 100-kHz/320-Vpp signal.

Introducing the 2013 JALA Ten.

Introducing the 2013 JALA Ten.

Determining the unique refractive index properties of solid polystyrene aerosol using broadband Mie scattering from optically trapped beads

A method is described to measure the refractive index dispersion with wavelength of optically trapped solid particles in air. Knowledge of the refraction properties of solid particles is critical for the study of aerosol; both in the laboratory and in the atmosphere for climate studies. Single micron-sized polystyrene beads were optically trapped in air using a vertically aligned counter-propagating configuration of focussed laser beams. Each bead was illuminated using white light from a broadband light emitting diode (LED) and elastic scattering within the bead was collected onto a spectrograph. The resulting Mie spectra were analysed to accurately determine polystyrene bead radii to ±0.4 nm and values of the refractive index to ±0.0005 over a wavelength range of 480-700 nm. We demonstrate that optical trapping combined with elastic scattering can be used to both accurately size polystyrene beads suspended in air and determine their wavelength dependent refractive index. The refractive index dispersions are in close agreement with reported values for polystyrene beads in aqueous dispersion. Our results also demonstrate a variation in the refractive index of polystyrene, from bead to bead, in a commercial sample. The measured variation highlights that care must be taken when using polystyrene beads as a calibration aerosol.

Air-Flow-Based Single-Cell Dispensing System

We discuss a design and fabrication approach to increase the success rate of single-cell dispensing. Two pairs of capacitance sensors are placed in a biochip to detect the flow velocity of cells and air pressure is applied to eject cells by synchronizing the timing. A comprehensive design theory, which takes into account the back-pressure caused by the air pressure, the response time of the system, the sensor properties and the delay of the dispensing from the air pressure, is developed in order to minimize the disturbance of the system and maximize the throughput of the ejection system. Then, the system theoretically has a capability to eject 3 cells/s and the maximum flow velocity is 10 mm/s. The novelty of the system is that the biochip is disposable, which is unlike the conventional mechanical inkjet system; because the biochip is low cost and disposable this prevents contamination and means the drive system is reusable. Finally, we succeeded in automatic dispensing of a single polystyrene bead (100 μm) from a biochip to a culture well atmosphere using the developed cell ejection system with a success rate of 50%. Furthermore, we also succeeded in single swine oocyte dispensing by using the developed system.

Mass sensors with mechanical traps for weighing single cells in different fluids

We present two methods by which single cells can be mechanically trapped and continuously monitored within the suspended microchannel resonator (SMR) mass sensor. Since the fluid surrounding the trapped cell can be quickly and completely replaced on demand, our methods are well suited for measuring changes in cell size and growth in response to drugs or other chemical stimuli. We validate our methods by measuring the density of single polystyrene beads and Saccharomyces cerevisiae yeast cells with a precision of approximately 10(-3) g cm(-3), and by monitoring the growth of single mouse lymphoblast cells before and after drug treatment.