Separation of Cells and Microparticles in Insulator-Based Electrokinetic Systems

Abstract

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 (t R,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 (t R,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 (R s), where R s 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.

Characterization of the Nonlinear Electrophoretic Behavior of Colloidal Particles in a Microfluidic Channel

“…Despite their similarities, there is still merit in distinguishing type 3 from type 1 microparticles. In most electrokinetic separation devices, , the electric field in the system will be in a range under which the effects of nonlinear electrophoresis would be significant for type 1 particles, while almost nonexistent for type 3 particles, as type 3 particles have a “delayed” response to nonlinear electrophoresis. This response is consistent with the lower surface charge of these particles in comparison to type 1 particles.…”

“…This response is consistent with the lower surface charge of these particles in comparison to type 1 particles. For practical purposes, when designing a microparticle separation, there is a clear distinction between type 1 and type 3 particles, where the former would clearly show in their migration the effects of nonlinear electrophoresis, while the latter would not. , These observations are not unprecedented, since Kumar et al reported a linear v EP of particles even at high fields’ strengths. Although the size and surface charge of the particles they employed are different from the ones used in our experiments, these being 4 μm in diameter and moderately charged particles, the reported particle movement indicated the characteristic type 3 behavior.…”

Dependence of Nonlinear Electrophoresis on Particle Size and Electrical Charge

“…New EK-based studies are continuously reported, as this is a rapidly growing field. Novel developments range from multiple frequency flows to two-phase flow applications, including hybrid systems that combine electrochemistry with EK. − Our group employs the data obtained in this study for the design of separation processes of particles and cells in microchannels that feature insulating posts which enhance nonlinear EK effects. , …”

“…19−22 Our group employs the data obtained in this study for the design of separation processes of particles and cells in microchannels that feature insulating posts which enhance nonlinear EK effects. 17,23 ■ THEORY EK phenomena are classified in terms of their dependence on the electric field. Linear EK phenomena, also called first-kind phenomena, depend on the permanent surface charge, while nonlinear phenomena, also called second-kind phenomena, depend on the bulk charge.…”

Fine-Tuning the Characteristic of the Applied Potential To Improve AC-iEK Separations of Microparticles

“…Microscale electrokinetics (EK) methods offer great potential for the manipulation and analysis of target bioparticlesincluding not only macromolecules but also nanoparticles and microparticles. These methods combine the attractive features of microfluidics (e.g., portability, low cost, and short response time) with the robustness and simplicity of EK phenomena to achieve label-free particle manipulation as function of the electric properties of particles and suspending solution. − EK-driven microfluidics have been successfully applied for the detection of DNA and proteins, − enrichment and assessment of cancer cells, − electrofusion of mammalian cells to create hybrid cells, and separation of microparticles and cells. − …”