Nanolocalized Single-Cell-Membrane Nanoelectroporation: For higher efficiency with high cell viability.
TL;DR: The process not only controls the precise delivery mechanism into the single cell with membrane reversibility but also provides spatial, temporal, and qualitative dosage control, which might be beneficial for therapeutic and biological cell studies.
Abstract: TODAY, SINGLE-CELL RESEARCH is of great interest to analyze cell-to-cell or cell-to-environment behavior with their intracellular compounds, where bulk measurements of millions of cells together can provide an average value. To deliver biomolecules in a precise and localized way into single living cells with a high transfection rate and high cell viability is a challenging and promisible task for biological and therapeutic research.
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TL;DR: In this review, this review of the recent advances in single-cell technologies and their applications insingle-cell manipulation, diagnosis, and therapeutics development are described.
Abstract: The investigation of human disease mechanisms is difficult due to the heterogeneity in gene expression and the physiological state of cells in a given population. In comparison to bulk cell measurements, single-cell measurement technologies can provide a better understanding of the interactions among molecules, organelles, cells, and the microenvironment, which can aid in the development of therapeutics and diagnostic tools. In recent years, single-cell technologies have become increasingly robust and accessible, although limitations exist. In this review, we describe the recent advances in single-cell technologies and their applications in single-cell manipulation, diagnosis, and therapeutics development.
63 citations
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.
41 citations
01 Jan 2020
TL;DR: This chapter mainly focuses on different physical drug-delivery techniques such as electroporation, optoporation, mechanopsoration, magnetoporation and hybrid techniques along with their working mechanisms, advantages, disadvantages, and limitations.
Abstract: Delivery of exogenous materials or cargo such as drugs, proteins, peptides, and nucleic acids into cells is a vital segment in molecular and cellular biology for potential cellular therapy and drug-discovery applications contributing toward personalization of medicine. Over the years, drug-delivery techniques have been developed in order to gain more control over the drug dosage, targeted delivery, and to minimize side effects. The major drug-delivery techniques can be classified as viral, chemical, and physical methods. Viral vectors are prominently used for gene therapy; however, they are cell-specific and have an immune response with high toxicity. Chemical methods are often limited by the low efficiency of plasmid delivery into different cell types due to plasmid degradation and toxicity. Considering these limitations, different physical methods such as photoporation, gene gun, hydrodynamic injection, electroporation, and mechanoporation, etc., are being widely developed for highly efficient cargo delivery with low toxicity. These methods are able to create transient hydrophilic membrane pores to deliver cargos into cells using different physical energies. Currently, ex vivo cargo delivery is widely studied while few in vivo applications have been developed. Concerning several obstacles to cargo delivery into cells, this chapter mainly focuses on different physical drug-delivery techniques such as electroporation, optoporation, mechanoporation, magnetoporation, and hybrid techniques along with their working mechanisms, advantages, disadvantages, and limitations. An insight into the future prospects and real-time applications of these techniques is also discussed.
18 citations
TL;DR: A space–time (x,y,t) multiphysics model based on quasi-static Maxwell’s equations and nonlinear Smoluchowski's equation has been developed to investigate the electroporation phenomenon induced by pulsed electric field in multicellular systems having irregularly shape.
Abstract: Electroporation technique is widely used in biotechnology and medicine for the transport of various molecules through the membranes of biological cells. Different mathematical models of electroporation have been proposed in the literature to study pore formation in plasma and nuclear membranes. These studies are mainly based on models using a single isolated cell with a canonical shape. In this work, a space–time (x,y,t) multiphysics model based on quasi-static Maxwell’s equations and nonlinear Smoluchowski’s equation has been developed to investigate the electroporation phenomenon induced by pulsed electric field in multicellular systems having irregularly shape. The dielectric dispersion of the cell compartments such as nuclear and plasmatic membranes, cytoplasm, nucleoplasm and external medium have been incorporated into the numerical algorithm, too. Moreover, the irregular cell shapes have been modeled by using the Gielis transformations.
10 citations
01 Jan 2016
TL;DR: Electroporation technique opens up the new window for the manipulation of genomics, proteomics, trascriptomics, metabolomics, or fluxomics at single cell level for biological research and therapeutic applications.
Abstract: Single-cell analysis is a powerful technique to understand cell to cell or cell to environment behaviors. It can provide detailed information of cell proliferation, differentiation, and different responses to external stimuli and intracellular reactions. For single cell analysis, electroporation or electropermeabilization is an efficient and fast method, where high external electric field is applied on cell membrane to form transient membrane pores to deliver ions or molecules in or out of the cell. Conventional electroporation or bulk electroporation (BEP) is performed in a batch mode with millions of cells in suspension, which can only provide an average value. Since the last decade, microfabricated devices have been developed to perform single cell electroporation, where electric field is only intense on a single cell, resulting in high transfection efficiency with high cell viability compared to BEP. Single cell analysis using electroporation technique can be performed with microfabricated electrode arrays, carbon fiber microelectrodes, micropipettes or electrolyte-filled capillaries-based devices. Recently developed nanofabricated electrodes can realize localized single cell electroporation for delivery of different molecules with high transfection efficiency and high cell viability. Thus, single cell electroporation technique opens up the new window for the manipulation of genomics, proteomics, trascriptomics, metabolomics, or fluxomics at single cell level. This chapter emphasizes the recent advancement of electroporation technique for cellular delivery and the analysis method which might be potentially applicable for biological research and therapeutic applications.
10 citations
References
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939 citations
"Nanolocalized Single-Cell-Membrane ..." refers background in this paper
...After the pulse is withdrawn, the membrane reseals again; this phenomenon is known as reversible electroporation [3]....
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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.
756 citations
"Nanolocalized Single-Cell-Membrane ..." refers background in this paper
...As the high electric field intensifies, the cell membrane creates transient membrane nanopores to deliver biomolecules from the outside to the inside of the cell [1], [2]....
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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.
444 citations
"Nanolocalized Single-Cell-Membrane ..." refers background in this paper
...A very high electric field can permanently deform the cell membrane, resulting in cell death, the process known as irreversible electroporation [4]....
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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.
296 citations
"Nanolocalized Single-Cell-Membrane ..." refers background in this paper
...A larger electrode can provide higher transfection with high cell viability using electrophoresis-driven ion transportation, where the voltage requirement is higher [10], [11]....
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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.
268 citations
"Nanolocalized Single-Cell-Membrane ..." refers background or methods in this paper
...A larger electrode can provide higher transfection with high cell viability using electrophoresis-driven ion transportation, where the voltage requirement is higher [10], [11]....
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...Recently, the electroporation technique has focused on localizing the membrane area of the single cell, where the electric field can focus more precisely in a local area of the single cell; this process is known as localized single-cell-membrane electroporation (LSCMEP) [8]–[11]....
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...Some literature had suggested that membrane nanopores can open up for several seconds to minutes [9], [11]....
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