Single-Cell Electroporation with Real-Time Impedance Assessment Using a Constriction Microchannel

Abstract

The electroporation system can serve as a tool for the intracellular delivery of foreign cargos. However, this technique is presently limited by the inaccurate electric field applied to the single cells and lack of a real-time electroporation metrics subsystem. Here, we reported a microfluidic system for precise and rapid single-cell electroporation and simultaneous impedance monitoring in a constriction microchannel. When single cells (A549) were continuously passing through the constriction microchannel, a localized high electric field was applied on the cell membrane, which resulted in highly efficient (up to 96.6%) electroporation. During a single cell entering the constriction channel, an abrupt impedance drop was noticed and demonstrated to be correlated with the occurrence of electroporation. Besides, while the cell was moving in the constriction channel, the stabilized impedance showed the capability to quantify the electroporation extent. The correspondence of the impedance variation and electroporation was validated by the intracellular delivery of the fluorescence indicator (propidium iodide). Based on the obtained results, this system is capable of precise control of electroporation and real-time, label-free impedance assessment, providing a potential tool for intracellular delivery and other biomedical applications.

Recent Advances in Microscale Electroporation

“…Examples of channel constrictions implemented for continuous EP. (A) Schematic of single-cell, continuous EP through a narrow uniform constriction (top) . COMSOL Multiphysics simulations at three different points in the channel demonstrate sufficiently high TMP, but EP is achieved only when cells were elongated in the channel.…”

“…COMSOL Multiphysics simulations at three different points in the channel demonstrate sufficiently high TMP, but EP is achieved only when cells were elongated in the channel. Reproduced from ref . Copyright 2020 MDPI under Creative Commons CC BY 4.0 license, .…”

“…Optimal EP conditions were determined by monitoring an impedance curve over time with a sharp descent, indicating maximum poration, followed by signal recovery, indicating high cell viability. Ye et al compared the impedance drop with fluorescent signals measured during single-cell continuous EP . The authors noted a ∼186 ms delay from the initial impedance drop until the earliest fluorescence detection.…”

Microfluidic and Nanofluidic Intracellular Delivery

“…Most microfluidic constriction-based intracellular delivery approaches [33,53,58,[61][62][63][64] provide high scalability, simplicity of operation, and cell type insensitive delivery with decent delivery efficiency. However, drawbacks such as channel clogging, inconsistent delivery due to cell size heterogeneity, low plasmid DNA transfection efficiency, and large cargo consumption should be addressed for wider use as a next-generation intracellular delivery method.…”

Single Cell Microwave Biosensor for Monitoring Cellular Response to Electrochemotherapy

“…Many researches are now focalizing on transposing electroporation studies "on-chip" [18]- [22] in order to avoid costly, time-consuming work and invasiveness of the routinely used methods, such as microscopy, flow cytometry and chemical tests. Although very effective, there is an ongoing trend to develop and employ new analysis methods [20], [23]-…”