Prednosti i nedostaci različitih pristupa generiranja impulsa za elektroporaciju

Electroporator is a generator of electric pulses that is used for permeabilization of cells. There are five major concepts of electroporation design. Capacitor discharge, square wave generator, and analog generator are used to generate classical electroporation pulses that are longer than microsecond and pulse forming network, and resonant charging generator that are used to generate nanosecond electroporation pulses. The choice of an electroporator design is always driven by the biotechnological or biomedical application. Electroporators can be used for introduction of small (electrochemotherapy) and large molecules (gene electrotransfer), cell fusion, insertion of proteins into cell membrane, electroporation of organelles, pasteurization, tissue ablation etc. Basic concepts and foreseeable future developments in electroporator design are presented in this article.Elektroporator je generator impulsa koji se koristi za permeabilizaciju stanica. Postoji pet glavnih izvedbi elektroporatora. Pražnjenje kondenzatora, generator pravokutnog valnog oblika i analogni generator se koriste za klasične elektroporacijske impulse koji su duži od mikrosekunde, a mreža za formiranje impulsa i generator s rezonantnim nabijanjem se primjenjuju za generiranje nanosekundnih elektroporacijskih impulsa. Izbor izvedebe elektroporatora vođen je uvijek biotehnološkom ili biomedicinskom primjenom. Elektroporatori se mogu koristiti za ubacivanje malih (elektrokemoterapija) i velikih molekula (elektro genski prijenos), fuziju stanica, umetanje proteina u staničnu membranu, elektroporaciju organela, pasterizaciju, ablaciju tkiva itd. U radu su prikazani temeljni pristupi u izvedbama elektroporatora i predvidivi budući razvoj

Cancer treatment: an overview of pulsed electric field utilization and generation

Patients diagnosed with cancer receive different types of treatments based on the type and the level of the tumour. An emerging treatment that differs from well-developed systematic therapies (i.e., Chemotherapy, Radiotherapy, and Immunotherapy) is Tumour Treating Field (TTF) treatment. Tumour behaviour under TTF treatment varies based on the electric field intensity; the process of exposing the tumour cells to an electric field is called electroporation. From the electrical perspective, the most efficient method for electroporation is to use a voltage pulse generator. Several pulse generator topologies have been introduced to overcome existing limitations, mitigate the drawbacks of classical generators, and provide more controllable, flexible, and portable solid-state voltage pulse generators. This paper provides a review of cancer treatment using TTF and highlights the key specifications required for efficient treatment. Additionally, potential voltage pulse generators are reviewed and compared in terms of their treatment efficacy and efficient use of electrical power

Optical injection of mammalian cells using a microfluidic platform

The use of a focused laser beam to create a sub-micron hole in the plasma membrane of a cell (photoporation), for the selective introduction of membrane impermeable substances (optical injection) including nucleic acids (optical transfection), is a powerful technique most commonly applied to treat single cells. However, particularly for femtosecond photoporation, these studies have been limited to low throughput, small-scale studies, because they require sequential dosing of individual cells. Herein, we describe a microfluidic photoporation system for increased throughput and automated optical injection of cells. Hydrodynamic focusing is employed to direct a flow of single-file cells through a focused femtosecond laser beam for photoporation. Upon traversing the beam, a number of transient pores potentially open across the extracellular membrane, which allows the uptake of the surrounding fluid media into the cytoplasm, also containing the chosen injection agent. The process is entirely automated and a rate of 1 cell/sec could readily be obtained, enabling several thousand cells to be injected per hour using this system. The efficiency of optically injecting propidium iodide into HEK293 mammalian cells was found to be 42 ± 8%, or 28 ± 4% taking into account the requirement of post-injection viability, as tested using Calcein AM. This work now opens the way for combining photoporation with microfluidic analyses, sorting, purification or on-chip cell culture studies

Cell separation using electric fields

The present invention involves methods and devices which enable discrete objects having a conducting inner core, surrounded by a dielectric membrane to be selectively inactivated by electric fields via irreversible breakdown of their dielectric membrane. One important application of the invention is in the selection, purification, and/or purging of desired or undesired biological cells from cell suspensions. According to the invention, electric fields can be utilized to selectively inactivate and render non-viable particular subpopulations of cells in a suspension, while not adversely affecting other desired subpopulations. According to the inventive methods, the cells can be selected on the basis of intrinsic or induced differences in a characteristic electroporation threshold, which can depend, for example, on a difference in cell size and/or critical dielectric membrane breakdown voltage. The invention enables effective cell separation without the need to employ undesirable exogenous agents, such as toxins or antibodies. The inventive method also enables relatively rapid cell separation involving a relatively low degree of trauma or modification to the selected, desired cells. The inventive method has a variety of potential applications in clinical medicine, research, etc., with two of the more important foreseeable applications being stem cell enrichment/isolation, and cancer cell purging