Nanolocalized single cell membrane nanoelectroporation
TL;DR: This report presents a nano-localized single cell nano-electroporation technique, where electroporation take place in a very precise and localized area on a single cell membrane to achieve high efficient delivery with high cell viability.
Abstract: © 2014 IEEE. Today single cell research is a great interest to analyze cell to cell or cell to environment behavior with their intracellular compounds, where bulk measurement can provide average value. To deliver biomolecules precise and localized way into single living cell with high transfection rate and high cell viability is a challenging and promisible task for biological and therapeutic research. In this report, we present a nano-localized single cell nano-electroporation technique, where electroporation take place in a very precise and localized area on a single cell membrane to achieve high efficient delivery with high cell viability. We fabricated 60nm gap with 40 nm triangular Indium Tin Oxide (ITO) based nano-eletcrode tip, which can intense electric field in a nano-localized area of a single cell to permeabilize cell membrane and deliver exogenous biomolecules from outside to inside of the cell. This device successfully deliver dyes, proteins into single cell with high cell viability (98%). The process not only control precise delivery mechanism into single cell with membrane reversibility, but also it can provide special, temporal and qualitative dosage control, which might be beneficial for therapeutic and biological cell studies.
TL;DR: This review article will emphasize the basic concept and working mechanism associated with electroporation, single cell Electroporation and biomolecular delivery using micro/nanofluidic devices, their fabrication, working principles and cellular analysis with their advantages, limitations, potential applications and future prospects.
Abstract: © 2018 IOP Publishing Ltd. The ability to deliver foreign molecules into a single living cell with high transfection efficiency and high cell viability is of great interest in cell biology for applications in therapeutic development, diagnostics and drug delivery towards personalized medicine. Many chemical and physical methods have been developed for cellular delivery, however most of these techniques are bulk approach, which are cell-specific and have low throughput delivery. On the other hand, electroporation is an efficient and fast method to deliver exogenous biomolecules such as DNA, RNA and oligonucleotides into target living cells with the advantages of easy operation, controllable electrical parameters and avoidance of toxicity. The rapid development of micro/nanofluidic technologies in the last two decades, enables us to focus an intense electric field on the targeted cell membrane to perform single cell micro-nano-electroporation with high throughput intracellular delivery, high transfection efficiency and cell viability. This review article will emphasize the basic concept and working mechanism associated with electroporation, single cell electroporation and biomolecular delivery using micro/nanoscale electroporation devices, their fabrication, working principles and cellular analysis with their advantages, limitations, potential applications and future prospects.
01 Jan 2021
01 Jan 2016
TL;DR: Using single-cell DNA devices to improve DNA delivery accuracy for retinoic acid-induced P19 neurons under optimal conditions and a motion model based on parameters from dynamic transport, including an anterograde state, a retrograde state, and a pausing state are described.
Abstract: Protein accumulation occurs in various neurodegenerative diseases, and one hypothesis to explain this phenomenon is based on defective axonal transport powered by molecular motors. Kinesin family motor proteins and the opposing dynein/dynactin motor complex proteins are the major motor proteins for transport along microtubules in axons. The interaction and regulation of transport via subunits of kinesin and the dynein/dynactin complex and how these subunits influence motor protein structure and their transport are not yet well understood. Notably, RNA interference (RNAi) knockdown is widely used to investigate motor behavior. However, RNAi knockdown results have varied. Therefore, we have used single-cell DNA devices to improve DNA delivery accuracy for retinoic acid-induced P19 neurons under optimal conditions. In addition, a mathematical method and physical hypothesis fitting with physical parameters have been used to build models. We describe a motion model based on parameters from dynamic transport, including an anterograde state, a retrograde state, and a pausing state. The residence time between two transition points shows the ability of a motion mode to maintain its state and resist the tendency to switch. Thus, the model could explain the characteristics and influences of each subunit.
TL;DR: In this article, the authors used a Coulter Counter with a hydrodynamic focusing orifice to measure the dielectric breakdown of human and bovine red blood cells in a homogeneous electric field between two flat platinum electrodes.
Abstract: With human and bovine red blood cells and Escherichia coli B, dielectric breakdown of cell membranes could be demonstrated using a Coulter Counter (AEG-Telefunken, Ulm, West Germany) with a hydrodynamic focusing orifice. In making measurements of the size distributions of red blood cells and bacteria versus increasing electric field strength and plotting the pulse heights versus the electric field strength, a sharp bend in the otherwise linear curve is observed due to the dielectric breakdown of the membranes. Solution of Laplace's equation for the electric field generated yields a value of about 1.6 V for the membrane potential at which dielectric breakdown occurs with modal volumes of red blood cells and bacteria. The same value is also calculated for red blood cells by applying the capacitor spring model of Crowley (1973. Biophys. J. 13:711). The corresponding electric field strength generated in the membrane at breakdown is of the order of 4 . 10(6) V/cm and, therefore, comparable with the breakdown voltages for bilayers of most oils. The critical detector voltage for breakdown depends on the volume of the cells. The volume-dependence predicted by Laplace theory with the assumption that the potential generated across the membrane is independent of volume, could be verified experimentally. Due to dielectric breakdown the red blood cells lose hemoglobin completely. This phenomenon was used to study dielectric breakdown of red blood cells in a homogeneous electric field between two flat platinum electrodes. The electric field was applied by discharging a high voltage storage capacitor via a spark gap. The calculated value of the membrane potential generated to produce dielectric breakdown in the homogeneous field is of the same order as found by means of the Coulter Counter. This indicates that mechanical rupture of the red blood cells by the hydrodynamic forces in the orifice of the Coulter Counter could also be excluded as a hemolysing mechanism. The detector voltage (or the electric field strength in the orifice) depends on the membrane composition (or the intrinsic membrane potential) as revealed by measuring the critical voltage in E. coli B harvested from the logarithmic and stationary growth phases. The critical detector voltage increased by about 30% for a given volume on reaching the stationary growth phase.
TL;DR: This is a brief introduction to the emerging field of irreversible electroporation in medicine, where certain electrical fields when applied across a cell can have as a sole effect the permeabilization of the cell membrane, presumable through the formation of nanoscale defects in the cell membranes.
Abstract: This is a brief introduction to the emerging field of irreversible electroporation in medicine. Certain electrical fields when applied across a cell can have as a sole effect the permeabilization of the cell membrane, presumable through the formation of nanoscale defects in the cell membrane. Sometimes this process leads to cell death, primarily when the electrical fields cause permanent permeabilization of the membrane and the consequent loss of cell homeostasis, in a process known as irreversible electroporation. This is an unusual mode of cell death that is not understood yet. While the phenomenon of irreversible electroporation may have been known for centuries it has become only recently rigorously considered in medicine for various applications of tissue ablation. A brief historical perspective of irreversible electroporation is presented and recent studies in the field are discussed.
TL;DR: It is shown that nanochannel electroporation can deliver precise amounts of a variety of transfection agents into living cells, and is expected to have high-throughput delivery applications.
Abstract: A new device made up of a nanochannel and two microchannels can deliver well-defined amounts of molecules directly into cells without affecting cell viability.
TL;DR: A simple nanoelectroporation platform is demonstrated to achieve highly efficient molecular delivery and high transfection yields with excellent uniformity and cell viability and to offer excellent spatial, temporal, and dose control for delivery.
Abstract: Nondestructive introduction of genes, proteins, and small molecules into mammalian cells with high efficiency is a challenging, yet critical, process. Here we demonstrate a simple nanoelectroporation platform to achieve highly efficient molecular delivery and high transfection yields with excellent uniformity and cell viability. The system is built on alumina nanostraws extending from a track-etched membrane, forming an array of hollow nanowires connected to an underlying microfluidic channel. Cellular engulfment of the nanostraws provides an intimate contact, significantly reducing the necessary electroporation voltage and increasing homogeneity over a large area. Biomolecule delivery is achieved by diffusion through the nanostraws and enhanced by electrophoresis during pulsing. The system was demonstrated to offer excellent spatial, temporal, and dose control for delivery, as well as providing high-yield cotransfection and sequential transfection.