Novel Parallelized Electroporation by Electrostatic Manipulation of a Water-in-Oil Droplet as a Microreactor (original) (raw)
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Biochemistry and Biophysics Reports
We have developed a gene transfection method called water-in-oil droplet electroporation (EP) that uses a dielectric oil and a liquid droplet containing live cells and exogenous DNA. When a cell suspension droplet is placed between a pair of electrodes, an intense DC electric field can induce droplet deformation, resulting in an instantaneous short circuit caused by the droplet elongating and contacting the two electrodes simultaneously. Small transient pores are generated in the cell membrane during the short, allowing the introduction of exogenous DNA into the cells. The droplet EP was characterized by varying the following experimental parameters: applied voltage, number of short circuits, type of medium (electric conductivity), concentration of exogenous DNA, and size of the droplet. In addition, the formation of transient pores in the cell membrane during droplet EP and the transfection efficiency were evaluated.
Lab Chip, 2014
is an important technology for various biological applications. The exogenous DNA is commonly delivered into cells by using a strong electrical field to form transient pores in cellular membranes. However, the high voltage required in this electroporation process may cause cell damage. In this study, a dielectrophoretically-assisted electroporation was developed by using light-activated virtual microelectrodes in a new microfluidic platform. The DNA electrotransfection used a low applied voltage and an alternating current to enable electroporation and transfection. Single or triple fluorescencecarrying plasmids were effectively transfected into various types of mammalian cells, and the fluorescent proteins were successfully expressed in live transfected cells. Moreover, the multi-triangle optical pattern that was projected onto a photoconductive layer to generate localized non-uniform virtual electric fields was found to have high transfection efficiency. The developed dielectrophoretically-assisted electroporation platform may provide a simpler system for gene transfection and could be widely applied in many biotechnological fields.
Electroporation-mediated gene delivery
Advances in genetics, 2015
Electroporation has been used extensively to transfer DNA to bacteria, yeast, and mammalian cells in culture for the past 30 years. Over this time, numerous advances have been made, from using fields to facilitate cell fusion, delivery of chemotherapeutic drugs to cells and tissues, and most importantly, gene and drug delivery in living tissues from rodents to man. Electroporation uses electrical fields to transiently destabilize the membrane allowing the entry of normally impermeable macromolecules into the cytoplasm. Surprisingly, at the appropriate field strengths, the application of these fields to tissues results in little, if any, damage or trauma. Indeed, electroporation has even been used successfully in human trials for gene delivery for the treatment of tumors and for vaccine development. Electroporation can lead to between 100 and 1000-fold increases in gene delivery and expression and can also increase both the distribution of cells taking up and expressing the DNA as we...
Electroporation microchips for continuous gene transfection
Sensors and Actuators B-chemical, 2001
Electroporation is a technique with which DNA molecules can be delivered into cells in a chamber using high electric ®eld pulses. Compared to the commercial equipment, the fabricated¯ow-type electroporation microchip overcomes the limit in the amount of target cells and the potential risk of using high voltage, which are the two drawbacks in current electroporation technology. The¯owing mechanism eliminates the temperature-rising effect found in a commercial sterile plastic cuvette during electroporation and thus can improve the survival rate of cells after electroporation.
Progress in neurobiology, 2010
Electroporation is a widely used technique for enhancing the efficiency of DNA delivery into cells. Application of electric pulses after local injection of DNA temporarily opens cell membranes and facilitates DNA uptake. Delivery of plasmid DNA by electroporation to alter gene expression in tissue has also been explored in vivo. This approach may constitute an alternative to viral gene transfer, or to transgenic or knock-out animals. Among the most frequently electroporated target tissues are skin, muscle, eye, and tumors. Moreover, different regions in the central nervous system (CNS), including the developing neural tube and the spinal cord, as well as prenatal and postnatal brain have been successfully electroporated. Here, we present a comprehensive review of the literature describing electroporation of the CNS with a focus on the adult brain. In addition, the mechanism of electroporation, different ways of delivering the electric pulses, and the risk of damaging the target tiss...
Research & Reviews: Journal of Pharmacy and Pharmaceutical Sciences, 2015
Nowadays, many researchers in the field of gene delivery are focused on develop methods to produce nanoparticles with physicochemical characteristics in reproducible, continuous and scalable process without requiring post processing steps. Another key factor is to transpose conditions from in vivo gene delivery into cells and tissue to in vitro. In this context, microfluidics technology is emerging to replace the traditional methods to produce nanoparticles, incorporate nucleic acids into them and transfect cells in vitro. Within microfluidics, droplet-based systems have been highlighted by some special parameters provided by picoliter compartments created using two immiscible liquids. Several micro/ nanoparticles used in gene delivery can be produced through dropletbased systems with low polydispersity index, such as liposomes, bio/ polymeric nanoparticles, metal nanoparticles, polymersomes, microgels. In the case of in vitro transfection, it is known that conventional procedure in...
Electroporation of micro-droplet encapsulated HeLa cells in oil phase
Electrophoresis, 2010
Electroporation (EP) is a method widely used to introduce foreign genes, drugs or dyes into cells by permeabilizing the plasma membrane with an external electric field. A variety of microfluidic EP devices have been reported so far. However, further integration of prior and posterior EP processes turns out to be very complicated, mainly due to the difficulty of developing an efficient method for precise manipulation of cells in microfluidics. In this study, by means of a T-junction structure within a delicate microfluidic device, we encapsulated HeLa cells in micro-droplet of poration medium in oil phase before EP, which has two advantages: (i) precise control of cell-encapsulating droplets in oil phase is much easier than the control of cell populations or individuals in aqueous buffers; (ii) this can minimize the electrochemical reactions on the electrodes. Finally, we successfully introduced fluorescent dyes into the micro-droplet encapsulated HeLa cells in oil phase. Our results reflected a novel way to realize the integrated biomicrofluidic system for EP.
Size Specific Transfection to Mammalian Cells by Micropillar Array Electroporation
Scientific reports, 2016
Electroporation serves as a promising non-viral gene delivery approach, while its current configuration carries several drawbacks associated with high-voltage electrical pulses and heterogeneous treatment on individual cells. Here we developed a new micropillar array electroporation (MAE) platform to advance the electroporation-based delivery of DNA and RNA probes into mammalian cells. By introducing well-patterned micropillar array texture on the electrode surface, the number of pillars each cell faces varies with its plasma membrane surface area, despite their large population and random locations. In this way, cell size specific electroporation is conveniently carried out, contributing to a 2.5~3 fold increase on plasmid DNA transfection and an additional 10-55% transgene knockdown with siRNA probes, respectively. The delivery efficiency varies with the number and size of micropillars as well as their pattern density. As MAE works like many single cell electroporation are carried...