Cell lysis electroporation

Cell lysis Mechanical methods pressure shearing, ultrasonic disintegration, bead-mill homogenizers Nonmechanical methods enzymatic lysis, osmotic lysis, freezing and thawing, detergent-based lysis and electroporation... 

Cell lysis under a high electric field is referred to as electroporation [6], Under these conditions, the cell membrane experiences dramatic changes in permeability to macromolecules. The main applications of the electroporation include the electrotransformation of cells and the electroporative gene transfer by the uptake of foreign DNA or RNA (in plants, animals, bacteria, and yeast). The electric field generates permeable microspores at the cell membrane, so that the nucleic acid can be introduced by electroosmosis or diffusion. 

Cell lysis on a chip has been carried out by several different approaches. Detergents have been used to lyse cells on a chip. Thermal lysing on a chip has been carried out by placing the cell in the sample reservoir and then raising the temperature of the chip [50], A practical approach for microchip applications is lysis by electroporation. Since fluids are moved around on chips by the application of an electric field, its use in cell lysis is an obvious choice. 

However, in certain cases, both the cell membrane and cytoplasm properties cannot be ignored, for example during electroporation [46] or cell lysis [47], where the resistance of the cell membrane and the capacitance of the cytoplasm vary widely. In this case, the complete equivalent circuit model should be used. For a single-shelled spherical particle this includes the resistance of the membrane and the capacitance of the cytoplasm [48]. 

Fig. 2 (a) Microelectroporation device for cell lysis, (b) Device at various steps of the fabrication process after metallization and electrode-mold formation (left) and after electroplating right). (c) Dielectrophoresis (DEP) effect observed in the flow channels top). Saw-tooth microelectrodes acting as a DEP device for focusing intracellular materials after electroporation bottom). Reproduced from [23] with permission... 

One of the main applications of microfluidic electroporation devices is cell lysis. In these devices, the mechanical (shear force) or electrical forces are applied to rapture the cell membrane and release its intercellular contents. 

The most important function of electroporation is transfection, i.e., inserting the biological nanosamples into the living cell. Cell viability and transfection rate are the two most important indicators to evaluate the functionality of these microfluidic electroporatiOTi devices. The applied electric pulse must be ctuitroUed carefully. While the nanosample insertitHi should be as successful as possible, the viability of the cell may not be affected by applied external electric pulse. Compared with the cell lysis, the cell transfection usually performs by q)plying lower external electric field. 

Lu H, Schmidt MA, Jensen MF (2005) A mierofluidic electroporation device for cell lysis. Lab Chip 5 23-29... 

Above some electroporation threshold, the transmembrane potential cannot be further increased, and can even decrease due to transport of ions across the membrane [91, 95]. The phenomenon of membrane electroporation can also be understood in terms of tension. If the total membrane tension exceeds the lysis tension c ys, the vesicle ruptures. This corresponds to building up a certain critical transmembrane potential, = Pc- According to Eqs. (7.3) and (7.4), this porahon potenhal Pc depends on the inihal membrane tension Co as previously reported [59, 89, 90, 96, 97]. The crihcal hansmembrane potenhal for cell membranes is about IV (e.g., [98, 99]). 

When a large electric field is applied across a cell, the transmembrane potential is disrupted and pores are formed on the surface of the membrane. This phenomenon is called electroporation and is often used for gene transfection. As conventionally implemented, the process is reversible, and when the electric field is terminated, the pores close. The phenomenon can also be used to cause permanent disruption of the membrane, effectively lysing the cell. There have been several reports (Ml the use of electric lysis techniques in micM ofluidic devices [9-11]. Of particular interest, fast lysis of individual cells ( 33 ms) by electric pulses for chemical cytometry was demonstrated in a micM ofluidic platform [12]. These extremely rapid lysis methods which minimize unwanted effects of slow lysis (that may bias the results) make these techniques favorable for protein analysis when compared to chemical lysis techniques. One drawback of electric lysis is that much of the cell membrane, subcellular structures, and the nucleus may remain intact and thus can clog the channel or adhere to the surface, affecting the separation and limiting the capacity for reuse. 

In addition to these various chemical treatment methods, a number of physical methods of cell disruption can be used in a chip-based system. These physical methods include osmotic shock, which occurs when cells are suspended in a hypotonic solution shearing and fracturing of cells walls and membranes using microfabricated needles or spherical particles (beads) application of an electric field that causes electroporation ultrasonication of the cell sample and thermal lysis. 

Irreversible electroporation results in a loss of turgor, a leakage of cytoplasmic content, and lysis (Rubinsky, 2010), whereas reversible permeabilization leads to the formation of conductive channels across the cell membrane, though the electrically insulating properties will recover within seconds (Glaser et al., 1988 Angersbach et al, 2000). 

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