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.     

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 (delta(chi)) required for complete particle separation was about 1.75 x 10(-5) [cgs], corresponding to about 10(9) labeling ions per particle in this study. The throughput was around 7.2 x 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.     

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.     

SPLITT Cell Analytical Separation of Silica Particles. Non-Specific Crossover Effects: Does the Shear-Induced Diffusion Play a Role?

The gravitational split-flow lateral transport thin fractionation is known to be a fast, simple, theoretically tractable and tunable tool for the binary separation of molecular or particulate samples into different dimensional fractions. This fractionation is performed in a so-called SPLITT cell and is due to the combined effect of the gravitational force field and the flow rates inside the separation channel. It is known that separation performance is strongly dependent on the flow rate conditions and feed flow concentration, however, to date, few studies have been conducted to investigate the effect non-specific crossover has on separation. The aim of this work is to establish whether diffusive processes stemming from hydrodynamic effects contribute in any way to the quality of separation. A silica sample of known granule size distribution was chosen for this study which has environmental applications.     

A microfabricated electrical SPLITT system

A growing need for methods to analyze and prepare monodisperse nanoparticles on an industrial scale exists and may be solved by the application of split flow thin fractionation (SPLITT) at the microscale. Microfluidic systems of this type have the ability to separate nanoparticles with high precision in a continuous manner. A miniaturized SPLITT system can be fabricated using standard microfabrication technologies, works in a continuous mode, and can be used as a sample preparation instrument in a micro-total-analysis-system (micro-TAS). In this paper, a miniaturized electrical SPLITT system, which separates particles continuously based on electrophoretic mobility, has been characterized. The advantages of miniaturization have been elucidated. The various aspects of the micro SPLITT system discussed in this paper can be broadly classified into: micro SPLITT system design, fluidics modeling to refine the splitter arrangements, and experimental characterization of the SPLITT system. The design of the micro SPLITT system has been elucidated focusing on the two designs that were implemented. Fluid modeling, used to arrive at a new SPLITT design, was done using a commercially available CFD package to investigate behavior of the fluid in the microchannel with various splitter arrangements. Testing was done with nanoparticles of varying diameter and electrophoretic mobilities to verify the modeling results and demonstrate functionality of the SPLITT system. Particles eluted from both outlets of the SPLITT system were characterized using AFM and SEM to verify the function of the system.     

Assessment of the distribution of pesticides on soil particle fractions in simulated irrigation run-off using centrifugal SPLITT fractionation and ELISA

This study was designed to measure the distribution of pesticides within the mobile phase of simulated irrigation run-off water, using centrifugal split-flow thin-channel (SPLITT) fractionation, a novel technique providing a gentle separation of natural sediment and suspended particles. Particular attention is paid to the extraction of pesticide residues for enzyme-linked immunosorbent assay (ELISA) analysis; ELISA was used because of the limited sample size.Centrifugal SPLITT fractionation combined laminar flow hydrodynamics and centrifugal sedimentation to obtain a continuous binary separation of suspended particles. The non-destructive technique allowed an accurate separation of particles into fractions with divisions at 0.5, 2 and 10 μm, with those above 25 μm being performed by wet sieving. ELISA was used to analyse the concentration of endosulfan and diuron for each fraction generated by the SPLITT technique.This data can be used to determine the role that particulate fines and colloidal fractions play in the transport of bound organic pollutants within the environment and to examine prospects for remediation on farms.     

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.     

基于磁泳的生物分离分析技术

利用磁场诱导的微粒运动即磁泳对磁响应性粒子进行精细分离,是近年来发展起来的选择性分离细胞和高分子量核酸的有效技术。本文在阐明磁泳分离原理的基础上,介绍了磁泳分离的分流薄层分级技术、四极磁场流动分离技术和微芯片上的自由流磁泳分离技术的装置构造、工作原理及其在生物分离分析中的应用。     

Quantitative SPLITT fractionation of lagoon sediments

In this work, SPLITT Fractionation (split flow thin cell) is used to sort hydrodynamically sedimented particles coming from the Sacca di Goro, a lagoon-like system close to the Po River delta (Italy). First the possibility of performing quantitative mass separations with a SPLITT cell apparatus was checked on a standard silica sample of known particle size distribution (PSD). Environmental sediment samples and relative SPLITT sub-fractions were subject to Inductive Coupled Plasma--Atomic Emission Spectroscopy (ICP-AES) characterization for the following elements: Al, Fe, Cd, Cr, Cu, Ni, Mn, Pb and Zn. The distribution of these metals by particle size fractions has been investigated. The accuracy of the entire separation procedure has been also evaluated.     

基于磁泳的生物分离分析技术

利用磁场诱导的微粒运动即磁泳对磁响应性粒子进行精细分离,是近年来发展起来的选择性分离细胞和高分子量核酸的有效技术。本文在阐明磁泳分离原理的基础上,介绍了磁泳分离的分流薄层分级技术、四极磁场流动分离技术和微芯片上的自由流磁泳分离技术的装置构造、工作原理及其在生物分离分析中的应用。     

Analytical SPLITT cell fractionation: Linearity and resolution study

In this paper the analytical SPLITT (split flow thin cell) procedure is used to characterize the percentage composition of micronic polydisperse particulate samples at a given cut‐off size. The linearity and resolution of the separation method have been tested using specifically prepared starch samples, in order to compare the analytical process with two continuous (preparative) SPLITT procedures. Linearity has been checked by injecting a series of suspensions (at different concentrations) under five different flow rate conditions. Retrieval factors F were evaluated to verify the relative amount of sample exiting the cell outlets. The effective resolution has been assessed by inspecting the SPLITT fractions with an optical microscope, counting the granules, and evaluating the percentage of granules of expected size. It has been found that the resolution is very good (around 90%) and independent of sample distribution. It is seen from the comparison that in the analytical SPLITT mode sample resolution is usually around 85–90% and it is significantly better than that of the continuous SPLITT modes, thus making the analytical mode valuable in characterizing polydisperse samples. The method was tested for the characterization of a commercial starch sample.     

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.     

Field-flow fractionation of magnetic particles in a cyclic magnetic field

Although magnetic field-flow fractionation (MgFFF) is emerging as a promising technique for characterizing magnetic particles, it still suffers from limitations such as low separation efficiency due to irreversible adsorption of magnetic particles on separation channel. Here we report a novel approach based on the use of a cyclic magnetic field to overcome the particle entrapment in MgFFF. This cyclic field is generated by rotating a magnet on the top of the spiral separation channel so that magnetic and opposing gravitational forces alternately act on the magnetic particles suspended in the fluid flow. As a result, the particles migrate transversely between the channel walls and their adsorption at internal channel surface is prevented due to short residence time which is controlled by the rotation frequency. With recycling of the catch-release process, the particles follow saw-tooth-like downstream migration trajectories and exit the separation channel at velocities corresponding to their sedimentation coefficients. A retention model has been developed on the basis of the combined effects of magnetic, gravitational fields and hydrodynamic flow on particle migration. Two types of core-shell structured magnetic microspheres with diameters of 6.04- and 9.40-μm were synthesized and used as standard particles to test the proposed retention theory under varying conditions. The retention ratios of these two types of particles were measured as a function of magnet rotation frequency, the gap between the magnet and separation channel, carrier flow rate, and sample loading. The data obtained confirm that optimum separation of magnetic particles with improved separation efficiency can be achieved by tuning rotation frequency, magnetic field gradient, and carrier flow rate. In view of the widespread applications of magnetic microspheres in separation of biological molecules, virus, and cells, this new method might be extended to separate magnetically labeled proteins or organisms for multiplex analyte identification and purification.     

Analytical methods for separating and isolating magnetic nanoparticles

Despite the large body of literature describing the synthesis of magnetic nanoparticles, few analytical tools are commonly used for their purification and analysis. Due to their unique physical and chemical properties, magnetic nanoparticles are appealing candidates for biomedical applications and analytical separations. Yet in the absence of methods for assessing and assuring their purity, the ultimate use of magnetic particles and heterostructures is likely to be limited. In this review, we summarize the separation techniques that have been initially used for this purpose. For magnetic nanoparticles, it is the use of an applied magnetic flux or field gradient that enables separations. Flow based techniques are combined with applied magnetic fields to give methods such as magnetic field flow fractionation and high gradient magnetic separation. Additional techniques have been explored for manipulating particles in microfluidic channels and in mesoporous membranes. Further development of these and new analytical tools for separation and analysis of colloidal particles is critically important to enable the practical use of these, particularly for medicinal purposes.     

Impact of non‐ideal analyte behavior on the separation of protein aggregates by asymmetric flow field‐flow fractionation

Asymmetric flow field‐flow fractionation is a valuable tool for the characterization of protein aggregates in biotechnology owing to its broad size range and unique separation principle. However, in practice asymmetric flow field‐flow fractionation is non‐trivial to use due to the major deviations from theory and the influence on separation by various factors that are not fully understood. Here, we report methods to assess the non‐ideal effects that influence asymmetric flow field‐flow fractionation separation and for the first time identify experimentally the main factors that impact it. Furthermore, we propose new approaches to minimize such non‐ideal behavior, showing that by adjusting the mobile phase composition (pH and ionic strength) the resolution of asymmetric flow field‐flow fractionation separation can be drastically improved. Additionally, we propose a best practice method for new proteins.     

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 m 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.     

重力场流分离系统中载液及流速条件对分离的影响

重力场流分离作为最简单的一种场流分离技术,常用于分离微米级颗粒。选择两种不同粒径(20 μ m和6 μ m)的聚苯乙烯(PS)颗粒作为样品,通过改变载液中叠氮化钠浓度、混合表面活性剂的比例及载液流速,利用自行设计生产的重力场流分离(gravitational flow field-flow fractionation, GrFFF)仪器,对颗粒混合样品进行分离,得到了相关谱图与数据,考察了这3种因素对分离效果(保留比(R)、塔板高度(H))的影响。结果表明:20 μ m PS颗粒的R值均大于6 μ m PS颗粒的R值,H值均小于6 μ m颗粒的H值;PS颗粒的R值与H值均随着载液中叠氮化钠浓度的增加而增加;但随着载液流速的增加,R值增加,H值减小。该研究为GrFFF系统的开发及应用提供了重要的参考价值。     

Methods for continuous flow fractionation of microparticles: Outlooks and fields of application

The development of new methods for fractionating particles of a different nature is becoming more important in solving some scientific and technological problems. This paper presents a brief review in the theory and practice of the most common techniques for microparticle fractionation (0.1–100 μm). These are dry and wet sieving, elutriation, sequential filtration, split-flow thin fractionation (SPLITT system), field-flow fractionation (FFF), membrane filtration, and capillary electrophoresis. Special attention is paid to the FFF technique, which offers a unique potential for the separation of different materials, from biopolymers and microorganisms to colloidal and solid particles, and the estimation of their physical properties. An alternative version of sedimentation FFF is described, namely, the fractionation of microparticles in rotating coiled columns. The main advantages and limitations of the methods are revealed and their outlooks and fields of applications are envisaged.     

Optimized hybrid dielectrophoretic microchip for separation of bioparticles

Point-of-care diagnostics requires a smart separation of particles and/or cells. In this work, the multiorifice fluid fractionation as a passive method and dielectrophoresis-based actuator as an active tool are combined to offer a new device for size-based particle separation. The main objective of the combination of these two well-established techniques is to improve the performance of the multiorifice fluid fractionation by taking advantage of dielectrophoresis-based actuator for separating particles. Initially, by using numerical simulations, the effect of using dielectrophoresis-based actuator in multiorifice fluid fractionation on the separation of particles was investigated, and the size of the device was optimized by 25% compared to a device without dielectrophoresis-based actuator. Also, adding dielectrophoresis-based actuator to multiorifice fluid fractionation can extend the range of flow rates needed for separation. In the absence of dielectrophoresis-based actuator, the separation took place only when the flow rate is 100 μL/min, in the presence of dielectrophoresis-based actuator (20 Vp-p), the separation happened in flow rates ranging from 70 to 120 μL/min.