The feasibility of using dielectrophoresis for isolation of glioblastoma subpopulations with increased stemness.

Cancer stem cells (CSCs) are aggressive subpopulations with increased stem-like properties. CSCs are usually resistant to most standard therapies and are responsible for tumor repropagation. Similar to normal stem cells, isolation of CSCs is challenging due to the lack of reliable markers. Antigen-based sorting of CSCs usually requires staining with multiple markers, making the experiments complicated, expensive, and sometimes unreliable. Here, we study the feasibility of using dielectrophoresis (DEP) for isolation of glioblastoma cells with increased stemness. We culture a glioblastoma cell line in the form of neurospheres as an in vitro model for glioblastoma stem cells. We demonstrate that spheroid forming cells have higher expression of stem cell marker, nestin. Next, we show that dielectric properties of neurospheres change as a result of changing culture conditions. Our results indicate that spheroid forming cells need higher voltages to experience the same DEP force magnitude compared to normal monolayer cultures of glioblastoma cell line. This study confirms the possibility of using DEP to isolate glioblastoma stem cells.

Separation of Macrophages and Fibroblasts Using Contactless Dielectrophoresis and a Novel ImageJ Macro.

Background: This study presents a label-free method of separating macrophages and fibroblasts, cell types critically associated with tumors. Materials and Methods: Contactless dielectrophoresis (DEP) devices were used to separate fibroblasts from macrophages by selectively trapping one population. An ImageJ macro was developed to determine the percentage of each population moving or stationary at a given point in time in a video. Results: At 350Vrms, 20 kHz, and 1.25 µL/min, more than 90% of fibroblasts were trapped while less than 20% of macrophages were trapped. Conclusions: Contactless DEP was used to study macrophage and fibroblast separation as a proof-of-concept study for separating cells in the tumor microenvironment. The associated ImageJ macro could be used in other microfluidic cell separation studies.

A feasibility study for enrichment of highly aggressive cancer subpopulations by their biophysical properties via dielectrophoresis enhanced with synergistic fluid flow.

A common problem with cancer treatment is the development of treatment resistance and tumor recurrence that result from treatments that kill most tumor cells yet leave behind aggressive cells to repopulate. Presented here is a microfluidic device that can be used to isolate tumor subpopulations to optimize treatment selection. Dielectrophoresis (DEP) is a phenomenon where particles are polarized by an electric field and move along the electric field gradient. Different cell subpopulations have different DEP responses depending on their bioelectrical phenotype, which, we hypothesize, correlate with aggressiveness. We have designed a microfluidic device in which a region containing posts locally distorts the electric field created by an AC voltage and forces cells toward the posts through DEP. This force is balanced with a simultaneous drag force from fluid motion that pulls cells away from the posts. We have shown that by adjusting the drag force, cells with aggressive phenotypes are influenced more by the DEP force and trap on posts while others flow through the chip unaffected. Utilizing single-cell trapping via cell-sized posts coupled with a drag-DEP force balance, we show that separation of similar cell subpopulations may be achieved, a result that was previously impossible with DEP alone. Separated subpopulations maintain high viability downstream, and remain in a native state, without fluorescent labeling. These cells can then be cultured to help select a therapy that kills aggressive subpopulations equally or better than the bulk of the tumor, mitigating resistance and recurrence.