Continuous-flow, electrically-triggered, single cell-level electroporation

Electroporation creates transient openings in the cell membrane, allowing for intracellular delivery of diagnostic and therapeutic substances. The degree of cell membrane permeability during electroporation plays a key role in regulating the size of the delivery payload as well as the overall cell viability. A microfluidic platform offers the ability to electroporate single cells with impedance detection of membrane permeabilization in a high-throughput, continuous-flow manner. We have developed a flow-based electroporation microdevice that automatically detects, electroporates, and monitors individual cells for changes in permeability and delivery. We are able to achieve the advantages of electrical monitoring of cell permeabilization, heretofore only achieved with trapped or static cells, while processing the cells in a continuous-flow environment. We demonstrate the analysis of membrane permeabilization on individual cells before and after electroporation in a continuous-flow environment, which dramatically increases throughput. We have confirmed cell membrane permeabilization by electrically measuring the changes in cell impedance from electroporation and by optically measuring the intracellular delivery of a fluorescent probe after systematically varying the electric field strength and duration and correlating the pulse parameters to cell viability. We find a dramatic change in cell impedance and propidium iodide (PI) uptake at a pulse strength threshold of 0.87 kV/cm applied for a duration of 1 ms or longer. The overall cell viability was found to vary in a dose dependent manner with lower viability observed with increasing electric field strength and pulse duration. Cell viability was greater than 83% for all cases except for the most aggressive pulse condition (1[Formula: see text]kV/cm for 5[Formula: see text]ms), where the viability dropped to 67.1%. These studies can assist in determining critical permeabilization and molecular delivery parameters while preserving viability.