Improving device design in insulator-based electrokinetic tertiary separations.

This study presents a methodology for designing effective insulator-based electrokinetic (iEK) systems for separating tertiary microparticle samples, which can be extended to more complex samples. First, 144 distinct iEK microchannel designs were built considering different shapes and arrangements of the insulating posts. Second, a mathematical model was developed with COMSOL software to predict the retention time of each particle type in the microchannel, this allowed identifying the best channel designs for two distinct types of separations: charge-based and sized-based. Third, the experimental charge-based and size-based separations of the tertiary microparticle mixtures were performed employing the improved designs identified with COMSOL modeling. The experimental results demonstrated successful separation in terms of separation resolution and good agreement with COMSOL predictions. The findings from this study show that the proposed method for device design, which combines mathematical modeling with varying post shape and post arrangement is an effective approach for identifying iEK systems capable of separating complex microparticle samples.

Switching Separation Migration Order by Switching Electrokinetic Regime in Electrokinetic Microsystems.

Analyte migration order is a major aspect in all migration-based analytical separations methods. Presented here is the manipulation of the migration order of microparticles in an insulator-based electrokinetic separation. Three distinct particle mixtures were studied: a binary mixture of particles with similar electrical charge and different sizes, and two tertiary mixtures of particles of distinct sizes. Each one of the particle mixtures was separated twice, the first separation was performed under low voltage (linear electrokinetic regime) and the second separation was performed under high voltage (nonlinear electrokinetic regime). Linear electrophoresis, which discriminates particles by charge, is the dominant electrokinetic effect in the linear regime; while nonlinear electrophoresis, which discriminates particles by size and shape, is the dominant electrokinetic effect in the nonlinear regime. The separation results obtained with the three particle mixtures illustrated that particle elution order can be changed by switching from the linear electrokinetic regime to the nonlinear electrokinetic regime. Also, in all cases, better separation performances in terms of separation resolution (Rs) were obtained by employing the nonlinear electrokinetic regime allowing nonlinear electrophoresis to be the discriminatory electrokinetic mechanism. These findings could be applied to analyze complex samples containing bioparticles of interest within the micron size range. This is the first report where particle elution order is altered in an iEK system.

Effect of cell shape on nonlinear electrophoresis migration.

This study contributes to the renewed interest in the study of nonlinear electrophoresis of colloidal particles. In this work the influence of cell shape on electrophoretic migration under the nonlinear regimes of moderate and strong field regimes was assessed. Four types of bacterial and yeast cells (one spherical, three non-spherical) were studied and their electrophoretic mobilities for the moderate and strong electric field magnitude regimes were estimated experimentally. The parameter of sphericity was employed to assess the effect cell shape on the nonlinear electrophoresis migration velocity and corresponding mobility under the two electric field magnitude regimes studied. As particle migration under nonlinear electrophoresis depends on particle size and shape, the results in terms of mobilities of nonlinear electrophoresis were presented as function of cell hydrodynamic diameter and sphericity. The results indicated that the magnitude of the mobilities of nonlinear electrophoresis for cells increase with increasing cell size and increase with increasing deviations from spherical shape, which is indicated by lower sphericity values. The results presented here are the very first assessment of the two types of mobilities of nonlinear electrophoresis of cells as a function of size and shape.

Dependence of Nonlinear Electrophoresis on Particle Size and Electrical Charge.

This study focuses on the dependence of nonlinear electrophoretic migration of particles on the particle size and particle electrical charge. This is the first report of the experimental assessment of the mobilities of the nonlinear electrophoretic velocity of colloidal polystyrene microparticles under two distinct electric field dependences. A total of nine distinct types of polystyrene microparticles of varying size and varying electrical charge were divided into two groups to study separately the effects of particle size and the effects of particle charge. The mobilities of the nonlinear electrophoretic velocity of each particle type were determined in both the cubic and 3/2 regimes (µEP,NL(3) and µEP,NL(3/2)). The results unveiled that both mobilities had similar relationships with particle size and charge. The magnitude of both µEP,NL(3) and µEP,NL(3/2) increased with increasing particle size and decreased with increasing magnitude of particle charge. However, the observed trends were not perfect as discussed in the Results and Discussion section but still provide valuable information. These findings will aid in the design of future size-based and charge-based separations of particles and microorganisms.

Separation of Cells and Microparticles in Insulator-Based Electrokinetic Systems.

Presented here is the first continuous separation of microparticles and cells of similar characteristics employing linear and nonlinear electrokinetic phenomena in an insulator-based electrokinetic (iEK) system. By utilizing devices with insulating features, which distort the electric field distribution, it is possible to combine linear and nonlinear EK phenomena, resulting in highly effective separation schemes that leverage the new advancements in nonlinear electrophoresis. This work combines mathematical modeling and experimentation to separate four distinct binary mixtures of particles and cells. A computational model with COMSOL Multiphysics was used to predict the retention times (tR,p) of the particles and cells in iEK devices. Then, the experimental separations were carried out using the conditions identified with the model, where the experimental retention time (tR,e) of the particles and cells was measured. A total of four distinct separations of binary mixtures were performed by increasing the level of difficulty. For the first separation, two types of polystyrene microparticles, selected to mimic Escherichia coli and Saccharomyces cerevisiae cells, were separated. By leveraging the knowledge gathered from the first separation, a mixture of cells of distinct domains and significant size differences, E. coli and S. cerevisiae, was successfully separated. The third separation also featured cells of different domains but closer in size: Bacillus cereus versus S. cerevisiae. The last separation included cells in the same domain and genus, B. cereus versus Bacillus subtilis. Separation results were evaluated in terms of number of plates (N) and separation resolution (Rs), where Rs values for all separations were above 1.5, illustrating complete separations. Experimental results were in agreement with modeling results in terms of retention times, with deviations in the 6-27% range, while the variation between repetitions was between 2 and 18%, demonstrating good reproducibility. This report is the first prediction of the retention time of cells in iEK systems.

High-Resolution Charge-Based Electrokinetic Separation of Almost Identical Microparticles.

Well-established techniques, e.g., chromatography and capillary electrophoresis, are available for separating nanosized particles, such as proteins. However, similar techniques for separating micron-sized particles are still needed. Insulator-based electrokinetic (iEK) systems can achieve efficient microparticle separations by combining linear and nonlinear EK phenomena. Of particular interest are charge-based separations, which could be employed for separating similar microorganisms, such as bacterial cells of the same size, same genus, or same strain. Several groups have reported charge-based separations of microparticles where a zeta potential difference of at least 40 mV between the microparticles was required. The present work pushes the limit of the discriminatory capabilities of iEK systems by reporting the charged-based separation of two microparticles of the same size (5.1 µm), same shape, same substrate material, and with a small difference in particle zeta potentials of only 3.6 mV, which is less than 10% of the difference in previous studies. By building an accurate COMSOL Multiphysics model, which correctly accounts for dielectrophoresis and electrophoresis of the second kind, it was possible to identify the conditions to achieve this challenging separation. Furthermore, the COMSOL model allowed predicting particle retention times (tR,p) which were compared with experimental values (tR,e). The separations results had excellent reproducibility in terms of tR,e with variations of only 9% and 11% between repetitions. These findings demonstrate that, by following a robust protocol that involves modeling and experimental work, it is possible to discriminate between highly similar particles, with much smaller differences in electrical charge than previously reported.

Microscale electrokinetic-based analysis of intact cells and viruses.

Miniaturized electrokinetic methods have proven to be robust platforms for the analysis and assessment of intact microorganisms, offering short response times and higher integration than their bench-scale counterparts. The present review article discusses three types of electrokinetic-based methodologies: electromigration or motion-based techniques, electrode-based electrokinetics, and insulator-based electrokinetics. The fundamentals of each type of methodology are discussed and relevant examples from recent reports are examined, to provide the reader with an overview of the state-of-the-art on the latest advancements on the analysis of intact cells and viruses with microscale electrokinetic techniques. The concluding remarks discuss the potential applications and future directions.