On-Chip Integration of Impedance Cytometry for Inline Optimization of Dielectrophoretic Separations on Multiple Cellular Biophysical Metrics

Microfluidic cell separation by dielectrophoresis, based on biophysical and electrical physiology metrics, is often optimized using on-chip fluorescence microscopy or off-chip flow cytometry of the separated fractions. However, these techniques require fluorescent reporters or stained samples that operate as end point assays, preventing the separated cell fractions from being utilized for longitudinal or transplantation studies. Single-cell impedance cytometry has a small footprint for facile integration toward label-free quantification of the separated fractions based on cell size, viability, and biophysical metrics. However, this is limited by low impedance signal-to-noise ratios in the low-conductivity media optimal for dielectrophoretic separation and by irreversible dielectrophoretic cell capture on impedance acquisition electrodes, while its single-cell resolution ability is limited by the high channel depths used to enhance sample throughput. Herein, using viscoelastic flows for elasto-inertial cell focusing over the channel depth, the throughput of dielectrophoretic separation is maintained, and bubble formation is avoided, while the downstream voltage for impedance cytometry can be maximized without irreversible cell capture to enhance impedance sensitivity in the dielectrophoretic separation media. This multichannel cytometry capability is integrated for automated optimization of the dielectrophoretic enrichment of live circulating tumor cells released into the suspension of pancreatic cancer cell cultures, using impedance metrics to monitor the separated fractions for feedback toward the selection of live cells within specific size ranges and with minimized transmembrane voltage-induced cell damage.