Split-flow Thin Research Articles

A dry film technology for the manufacturing of 3-D multi-layered microstructures and buried channels for lab-on-chip

The development of innovative and reliable techniques for devices miniaturization are enabling the massive growth of lab on chip (LOC) applications. In this article, we briefly review the technological options for LOC microfabrication, then we present the optimization of a process for the realization of tridimensional multi-layered structures and buried channels in a microfluidic network using a photo-patternable dry film, with a potential for LOC manufacturing. The tuning of all the fabrication parameters is widely discussed and micrographs and optical profiler images are reported to show fabrication results. The fabrication process is used for a Split-flow-thin (SPLITT) fractionation cell configuration. SPLITT is a particle fractionation technique based on the combined effect of two laminar streams (the sample containing the particles and a carrier) flowing inside a thin microchannel and the action of a vertical driving force for particle displacement. Since the SPLITT implemented in this work is electrically driven, patterned electrodes (thickness: 100 nm) are also integrated in the flow cell walls. The functionality of the cell was tested first verifying the presence of proper flow conditions for microfluidic SPLITT (absence of mixing between the streams) and then proving electrical fractionation with two different proteins (BSA and β-lactoglobulin) at different levels of ionic strength. The flow of the streams within the microfluidic channel was also simulated by a numerical 2-D model exactly reproducing the cell geometry, with a good accordance with experimental results.

Split-Flow Thin (SPLITT) Cell Separations Operating under Sink-Float Mode Using Centrifugal and Gravitational Fields

Particles of two different densities can be continuously and rapidly separated by employing sink-float phenomenon in split-flow thin (SPLITT) cells using both centrifugal and gravitational fields. Separation of a binary mixture of submicron size latex beads differing in densities is achieved by feeding them as a suspension in a carrier liquid of intermediate density into a SPLITT cell subject to a centrifugal field. A steady state condition is rapidly achieved, and two particle types accumulate at opposing channel walls as they are carried by the laminar channel flow to two separate outlets. Similarly, micron size particles of two different densities are separated using a planar channel and the gravitational field. Equations are developed to predict throughput in terms of the number fractions of the separated particles determined by scanning electron microscopy. Experimental results from three SPLITT cell systems under different field and flow conditions are compared with theoretical predictions.

Separation of magnetic beads in a hybrid continuous flow microfluidic device

Magnetic separation of biological entities in microfluidic environment is a key task for a large number of bio-analytical protocols. In magnetophoretic separation, biochemically functionalized magnetic beads are allowed to bind selectively to target analytes, which are then separated from the background stream using a suitably imposed magnetic field. Here we present a numerical study, characterizing the performance of a magnetophoretic hybrid microfluidic device having two inlets and three outlets for immunomagnetic isolation of three different species from a continuous flow. The hybrid device works on the principle of split-flow thin (SPLITT) fractionation and field flow fractionation (FFF) mechanisms. Transport of the magnetic particles in the microchannel has been predicted following an Eulerian-Lagrangian model and using an in-house numerical code. Influence of the salient geometrical parameters on the performance of the separator is studied by characterizing the particle trajectories and their capture and separation indices. Finally, optimum channel geometry is identified that yields the maximum capture efficiency and separation index.

Operating regimes of a magnetic split-flow thin (SPLITT) fractionation microfluidic device for immunomagnetic separation

Magnetic bead-based immunoassays in the microfluidic format have attracted particular interest as it has several advantages over other microfluidic separation techniques. Magnetic split-flow thin fractionation (SPLITT) is a compact version of microfluidic sorting where a bidispersed or polydispersed suspension of magnetic particle–analyte conjugates can be selectively isolated into co-flowing streams of nearly monodispersed particles. Although the device offers capability of identifying and separating more than one target analytes simultaneously, its performance is sensitive to the slightest variation of the operating condition. Herein, we have numerically investigated the performance of a microscale magnetic SPLITT device. Using a coupled Eulerian–Lagrangian approach, we have evaluated the capture efficiency (CE) and separation index (SI) for each particle type collected at their designated outlet of the SPLITT device and identified the best regimes of operating parameters. While the CE figures are found to be best represented by a group variable Π, the SI values are better represented as function of the product of the group variables γ and β; the SI versus Π plots clearly separate into two basic trends: one for constant β (i.e., varying γ) and the other for constant γ (i.e., varying β). Our study prescribes the desired operating regimes of a microfluidic magnetophoretic SPLITT device in a practical immunomagnetic separation application.

Analytical and preparative applications of magnetic split-flow thin fractionation on several ion-labeled red blood cells

BackgroundMagnetic Split-flow thin (SPLITT) fractionation is a newly developed technique for separating magnetically susceptible particles. Particles with different field-induced velocities can be separated into two fractions by adjusting applied magnetic forces and flow-rates at inlets and outlets.MethodsMagnetic particles, Dynabeads, were used to test this new approach of field-induced velocity for susceptibility determination using magnetic SF at different magnetic field intensities. Reference measurements of magnetic susceptibility were made using a superconducting quantum interference device (SQUID) magnetometer. Various ion-labeled red blood cells (RBC) were used to study susceptibility determination and throughput parameters for analytical and preparative applications of magnetic SPLITT fractionation (SF), respectively. Throughputs were studied at different sample concentrations, magnetic field intensities, and channel flow-rates.ResultsThe susceptibilities of Dynabeads determined by SPLITT fractionation (SF) were consistent with those of reference measurement using a superconducting quantum interference device (SQUID) magnetometer. Determined susceptibilities of ion-labeled RBC were consistent within 9.6% variations at two magnetic intensities and different flow-rates. The determined susceptibilities differed by 10% from referenced measurements. The minimum difference in magnetic susceptibility required for complete separation was about 5.0 × 10-6 [cgs]. Sample recoveries were higher than 92%. The throughput of magnetic SF was approximately 1.8 g/h using our experimental setup.ConclusionMagnetic SF can provide simple and economical determination of particle susceptibility. This technique also has great potential for cell separation and related analysis. Continuous separations of ion-labeled RBC using magnetic SF were successful over 4 hours. The throughput was increased by 18 folds versus early study. Sample recoveries were 93.1 ± 1.8% in triplicate experiments.

Hollow-fiber flow/hyperlayer field-flow fractionation for the size characterization of airborne particle fractions obtained by SPLITT fractionation

Hollow-fiber flow field-flow fractionation (HF FlFFF) was applied for the separation and size characterization of airborne particles which were collected in a municipal area and prefractionated into four different-diameter intervals >5.0, 2.5-5.0, 1.5-2.5, <1.5 microm) by continuous split-flow thin (SPLIIT) fractionation. Experiments demonstrated the possibility of utilizing a hollow-fiber module for the high-performance separation of supramicron-sized airborne particles at steric/hyperlayer operating mode of HF FlFFF. Eluting particles during HF FlFFF separation were collected at short time intervals (approximately 10 s) for the microscopic examination. It showed that particle size and size distributions of all SPLITT fractions of airborne particles can be readily obtained using a calibration and that HF FlFFF can be utilized for the size confirmation of the sorted particle fraction during SPLITT fractionation.

Pinched inlet gravitational split-flow thin fractionation of airborne particles and analysis of size dependent level of PCDD/Fs

Fine particles in air have a direct influence on human health because they carry toxic chemicals that can be deposited in the human lung when inhaled. Thus, particle size distribution and size dependent level of contamination of the airborne particles are important parameters for the study and assessment of environmental pollution. In this study, gravitational split-flow thin (SPLITT) fractionation (or GSF), a semi-preparative scale separation technique for particles, was applied for the continuous size sorting of airborne particles collected in urban area. About 2.0 g of airborne particles was fractionated into four different size intervals (<1.5, 1.5-2.5, 2.5-5.0, and >5.0 microm), and the collected fractions were examined by electron microscopy for particle size distribution and analyzed for the size dependent levels of polychlorinated dibenzo-p-dioxins and furans (PCDD/Fs). It was found that more than 60% of particles including dissolved matters in weight were smaller than 5.0 microm and they contained more than 86% of the total PCDD/Fs amount in airborne particles.

Combination of gravitational SPLITT fractionation and field-flow fractionation for size-sorting and characterization of sea sediment

A combination of gravitational split-flow thin (SPLITT) fractionation and sedimentation/steric field-flow fractionation (Sd/StFFF) has been used for continuous size-sorting of a sediment sample and for size analysis of the collected fractions. An IAEA (International Atomic Energy Agency) sediment material was separated into four size fractions (with theoretical size ranges <1.0, 1.0-3.0, 3.0-5.0, and >5.0 microm in diameter) by means of a three-step gravitational SPLITT fractionation (GSF) for which the same GSF channel was used throughout. The GSF fractions were collected and examined by optical microscopy (OM) and by Sd/St FFF. The mean diameters of the GSF fractions measured by OM were within the size interval predicted by GSF theory, despite the theory assuming that all particles are spherical, which is not true for the sediment particles. The Sd/St FFF results showed that retention shifted toward shorter elution time (or larger size) than expected, probably because of the shape effect. The results from GSF, OM, and Sd/StFFF are discussed in detail.

Effect of flow development region and fringing magnetic force field on annular split-flow thin fractionation

Split-flow thin (SPLITT) fractionation devices have been widely used to separate macromolecules, colloids, cells and particles. Recently, the quadrupole magnetic flow sorter (QMS) has been reported in the literature as another family of SPLITT fractionation device. However, the separation performance observed in the experimental measurements is generally found to deviate from the ideal behaviour. Possible causes such as hydrodynamic lift force, high particle concentration and imperfect geometries have been extensively examined. However, the effects of flow development regions and fringing magnetic force field at the separation channel inlet and outlet, which are ignored by the theory, have not been investigated. The error introduced by ignoring these effects need to be rigorously studied so that the theory can be used to optimise operation flow rates with confidence. Indeed, we find in this paper that these ignored effects are responsible to the discrepancy between the experimental data and the theoretical predictions. A new theory has been proposed for optimisation of device operation.

Pinched inlet split flow thin fractionation for continuous particle fractionation: application to marine sediments for size-dependent analysis of PCDD/Fs and metals.

It is demonstrated that split-flow thin (SPLITT) fractionation, a continuous separation technique for sorting particles or macromolecules, can be utilized for the fractionation of environmental particles to study a size-dependent analysis of pollutants. In this study, focuses are made on the use of a pinched inlet gravitational SPLITT fractionation, a modified form of SPLITT channel formed by reducing the sample inlet thickness of the channel to improve separation efficiency, to separate marine sediments into five different sizes (<1.0, approximately 1.0 to 2.5, approximately 2.5 to 5.0, approximately 5.0 to 10, and approximately 10 to 53 microm). The resulting size fractions are examined with high resolution gas chromatography/high resolution mass spectrometry to determine the size-dependent distribution of polychlorinated dibenzo-p-dioxins and dibenzofurans along with a statistical data treatment and are analyzed with inductively coupled plasma-atomic emission spectrometry and graphite furnace atomic absorption spectrometry to ascertain its major and trace metals. It is shown that the combined analytical methods detailed in this study can be powerfully utilized in such a way as to analyze pollutant distribution and its concentration with regard to particle sizes for an environmental assessment.

Splitter Imperfections in Annular Split-Flow Thin Separation Channels: Experimental Study of Nonspecific Crossover

The separation performance of a split-flow thin (SPLITT) separation device depends on uniformity of channel thickness and the precise placement of the flow splitters at fixed distances between the channel walls. The observation of nonspecific crossover, that is, the transport of sample materials across the channel thickness without the influence of an applied field, has routinely been taken to indicate the presence of irregularities in splitter shape or placement. Computational fluid dynamics software may be used to predict the influence of splitter imperfections on nonspecific crossover, where it is assumed that sample transport is by convection alone. A previous study has shown how small inlet splitter imperfections can account for the relatively low levels of nonspecific crossover observed with typical annular SPLITT devices. This study, however, could not distinguish between the possible sources of nonspecific crossover; hydrodynamic lift or shear-induced diffusion could have contributed. To confirm the validity of the computational approach, a series of experiments has been carried out on a channel having a deliberately and severely bent splitter. Nonspecific crossover was measured for a range of inlet and outlet flow rate ratios, with the bent splitter placed at both the channel inlet and outlet. The severity of the splitter distortion was sufficient to produce significant nonspecific crossover over a wide range of flow conditions. Good agreement was found between experiment and prediction based on computational fluid dynamics, with experiment generally showing only slightly higher crossover than prediction. The quantitative agreement for this extreme case suggests that the contribution to nonspecific crossover due to geometrical imperfections can be well described using computational fluid dynamics.

Development of magnetic split-flow thin fractionation for continuous particle separation

Split-flow thin (SPLITT) fractionation is designed for preparative separation of macromolecules, colloids, and particles with throughputs typically in the gram(s) or less per hour range depending on the applied field. Magnetic separation using permanent magnets is fast, simple, highly selective, and economical. Magnetic SPLITT fractionation (SF) is a newly developed family of SF techniques for separating magnetically susceptible particles. Particles with different degrees of magnetic susceptibility can be separated into two fractions by adjusting perpendicularly applied magnetic forces and flow rates at inlets and outlets. The preparative applications of magnetic SF using various magnetically susceptible particles and mixtures were studied. Throughputs at various channel dimensions, sample concentrations, magnetic field strengths, and channel flow rates were examined. The minimum difference in magnetic susceptibility required for complete separation was about 3×10 −6 cgs. Sample recoveries were higher than 94%. Continuous particle separations using magnetic SF were successful over 8 h. The throughput of magnetic SF was approximately 3 g/h using our experimental setup. The throughput can be scaled up by using longer channel lengths, broader channel breadths, and stronger magnetic fields. This technique has shown good potential for becoming useful in separating magnetically susceptible colloids, and particles.

General theory for flow optimisation of split-flow thin fractionation

Recently, magnetic split-flow thin (SPLITT) fractionation has been developed to separate macromolecules, colloids, cells and particles. However, the previous theory, developed for an infinitely long channel, needs to be improved to consider the flow transit regimes at both inlet and outlet. In this paper, we describe a new approach to optimising flow-rates for particle separation which considers the effect of flow transit region. Surprisingly, the critical particle migration velocities derived by the present theory are identical to the previous simplified theory. Therefore, the previous simplified theory may have wider application than might have been expected. As a test of our theory, a numerical simulation based on solving Navier–Stokes equations has also been carried out for a magnetic SPLITT device. The trajectory of a particle with the critical migration velocity is exactly as expected by our theory. Following experimental validation, this work will facilitate the design of new SPLITT fractionation systems with smaller aspect ratio.

Splitter imperfections in annular split-flow thin separation channels: effect on nonspecific crossover.

The separation performance of split-flow thin (SPLITT) separation channels generally falls short of ideal behavior. There are many possible contributing factors to the loss of separation resolution, and these are discussed in the text. The possibility that small imperfections in the splitters play a significant role is examined in this study. Computational fluid dynamics is used to determine the flow pattern within an annular SPLITT channel having small imperfections in the inlet splitter. These results are used to calculate the nonspecific crossover of particles from the inner annular inlet to the outer annular outlet under various flow rate regimes. Nonspecific crossover, obtained through convective transport alone, and not the result of field-induced transport, is often used as a check of channel behavior. The results of a typical experimental determination of nonspecific crossover are included for comparison. It is concluded that geometrical imperfections can indeed play a significant role in the loss of resolution observed for these systems.

Particle magnetic susceptibility determination using analytical split-flow thin fractionation

Magnetic split-flow thin (SPLITT) fractionation is a newly developed SPLITT fractionation subtechnique for separating magnetically susceptible particles. Here, we describe determination of particle magnetic susceptibilities using a novel method, analytical magnetic SPLITT fractionation. This method is based on particle retrieval calculation under known flow-rates and magnetic field strengths. The flow axis of separation channel was oriented parallel to gravity to exclude gravitational effects on separation. Various carrier magnetic susceptibilities, magnet interpolar gap widths, and ion-labeled particles were studied experimentally, resulting in successful determination of magnetic susceptibilities for magnetically susceptible particles and various ion-labeled particles. The minimum level of particle magnetic susceptibility ( χ) that could be determined in this study was about 5.0·10 −7 (cgs). The minimum number of erbium labeling ions per particle required for silica susceptibility determination was found to be 1.57·10 10. These determined values were consistent with relative standard deviations within 8% variation and within 9% of standard reference measurements using a superconducting quantum interference device magnometer. Analytical magnetic SPLITT fractionation shows good potential for use in simple determination of particle magnetic susceptibilities.

On-line particle concentrator with upstream ultrafiltration in continuous SPLITT fractionation.

An upstream ultrafiltration (UU) method is employed for the on-line concentration of collected particle fraction and for the convenient regulation of flow rates in split-flow thin (SPLITT) fractionation. Concentration of the collected particle fraction is necessary to fractionate particles by smaller cutoff diameters in SPLITT fractionation. By introducing a simple device utilizing upstream ultrafiltration with tangential flow, particle solution collected at each SP

Study of magnetic particles pulse-injected into an annular SPLITT-like channel inside a quadrupole magnetic field

Advantages of the continuous magnetic flow sorting for biomedical applications over current, batch-wise magnetic separations include high throughput and a potential for scale-up operations. A continuous magnetic sorting process has been developed based on the quadrupole magnetic field centered on an annular flow channel. The performance of the sorter has been described using the conceptual framework of split-flow thin (SPLITT) fractionation, a derivative of field-flow fractionation (FFF). To eliminate the variability inherent in working with a heterogenous cell population, we developed a set of monodisperse magnetic microspheres of a characteristic magnetization, and a magnetophoretic mobility, similar to those of the cells labeled with a magnetic colloid. The theory of the magnetic sorting process has been tested by injecting a suspension of the magnetic beads into the carrier fluid flowing through the sorter and by comparing the theoretical and experimental recovery versus total flow-rate profiles. The position of the recovery maxima along the total flow-rate axis was a function of the average bead magnetophoretic mobility and the magnetic field intensity. The theory has correctly predicted the position of the peak maxima on the total flow-rate axis and the dependence on the bead mobility and the field intensity, but has not correctly predicted the peak heights. The differences between the calculated and the measured peak heights were a function of the total flow-rate through the system, indicating a fluid-mechanical origin of the deviations from the theory (such as expected of the lift force effects in the system). The well-controlled elution studies using the monodisperse magnetic beads, and the SPLITT theory, provided us with a firm basis for the future sorter evaluation using cell mixtures.

Continuous split-flow thin cell and gravitational field-flow fractionation of wheat starch particles

The combined employment of the SPLITT (split-flow thin) cell — a relatively new system for fast, continuous binary separation — and of gravitational field-flow fractionation (GrFFF) — a fractionation technique suitable for micron particle size distribution determination — was investigated for starch separation and characterization. Emphasis is placed on the main advantages of both techniques: operating under gentle earth gravity field, low cost and ease of maintenance. The reproducibility of GrFFF is demonstrated. Both the SPLITT separation and GrFFF fractionation results were checked by optical microscopy. Application examples of typical starch fractionation experiments are reported and discussed.

Size–elemental characterization of suspended particle matter by split-flow thin cell fractionation and slurry analysis–electrothermal atomic absorption spectrometry

An analytical methodology is presented for characterizing SPM (suspended particle matter) from natural waters by using the coupling between the SPLITT (split-flow thin) cell separation and elemental spectroscopic identification. The results of a systematic study validating direct slurry elemental analysis by ETAAS (electrothermal atomic absorption spectrometry) are reported for the determination of Al and Fe in mineral particulates, considered representative models of real SPM. Slurry analysis of clay-like particle materials was accurate in the particle size range up to 25 μm, provided that slurry concentrations met quantitative sampling conditions. The coupled SPLITT–ETAAS technique was then evaluated for the dimensional and elemental characterization of the same samples. The developed procedure was applied to a real SPM sample from the Po river, where trace elements of environmental interest were determined for the whole fraction in the 0.2–25 μm dimensional range as well as for some sub-fractions obtained by SPLITT fractionation.

Magnetic split-flow thin fractionation of magnetically susceptible particles

We recently built a magnetic separation system to extend the applications of split-flow thin (SPLITT) fractionation to magnetically susceptible particles. Here, we characterize the magnetic SPLITT system using magnetically susceptible particles and ion-labeled particles. The flow axis of separation channel was orientated parallel and perpendicular to gravitational forces to exclude and include, respectively, gravitational effects on separation. Both operating modes were used to test the theory experimentally, with emphasis on the parallel mode. The magnetic susceptibilities of carrier and ion-labeled particles were varied, and various ion-labeled and unlabeled particles were studied experimentally, resulting in successful separation of labeled particles, yeasts, and cells from unlabeled ones. The minimal difference in magnetic susceptibility (Δ χ) required for complete particle separation was about 1.75×10 −5 [cgs], corresponding to about 10 9 labeling ions per particle in this study. The throughput was around 7.2×10 8 particles/h using the present setup. Magnetic SPLITT fractionation shows good potential for use in obtaining particles magnetic susceptibilities from a simple theoretical treatment.