**3.2. Pore formation on SCEP**

Chen et al. demonstrated localized single cell membrane electroporation (LSCMEP) by using microfluidic device. Where ITO thin film was used as microelectrode with 1 µm gap between two micro-electrodes. The ITO microelectrode with 100 nm thickness and 2 µm width intense electric field much more in between two microelectrode gap [63]. Fig.5. shows the device

**Figure 5.** Fabrication process of ITO microelectrode based localized single cell electroporation chip. (a) Fabrication process step (b) Optical microscope image of patterned ITO microelectrodes. (c) SEM image of ITO microelectrodes

with micro channel (FIB etch), Permission to reprint obtained from Springer [63].

fabrication for localized electroporation experiment.

70 Advances in Micro/Nano Electromechanical Systems and Fabrication Technologies

In single cell electroporation technique, electroporation occurs in adherent cell and tissue. However single cell electroporation can be visualized for cell in suspension. In BEP, mostly the cells are in suspension as spheres, in which homogeneous electric field can be applied. But for single cell electroporation, electric field is in inhomogeneous form, which targets on a particular cell without effecting neighboring cells. Generally cell membrane described in terms of fluid mosaic membrane model [64]. Due to application of an electric electric field, the formation of pores into the cell membrane depends upon field strength with low conductance, which is approximated as electrical capacitors with infinite resistance. The pore as liquid capacitor which converts to the electrical force associated with transmembrane potential U into an expanding pressure within the aqueous pore interior [65-68]. The pore creation energy ΔE can be calculated with pressure balance by removal of planar area πr<sup>2</sup> and creation of a cylindrical pore edge of length 2πr, can be written as

$$
\Delta E = 2\Pi \gamma r - \Pi r^2 \Gamma \tag{5}
$$

where surface energy approximately Г= 1× 10 J/m<sup>2</sup> and the edge energy approximately Υ= 1 to 6 × 10-11 J/m [69-71]. Here Υ is constant even it is a function of r [70, 72-73]. To expand the pore radius from zero radious to r can be written as

$$
\Delta E(r) = 2\Pi \gamma r - \Pi r^2 \Gamma + A \,/\, r^4 \tag{6}
$$

The first term is energy related stressed pore edge with length 2πr. The second term is energy to remove a circular flat lipid membrane having energy per unit area Г and the third term is steric repulsion of the lipids with constant A. Fourth term arises when transmembrane potential Vm is nonzero, which is related to,

$$-0.5 \text{C}\_p V\_m^2 r^2 \tag{7}$$

The transmembrane potential ΔΨE, in a uniform electric field E at a point M with time t can be written as

$$
\Delta\boldsymbol{\psi}\_E(t) = \boldsymbol{\psi}\_{in} - \boldsymbol{\psi}\_{out} = -f\mathbf{g}(\boldsymbol{\lambda})\boldsymbol{RE}\cos\theta(\boldsymbol{M}) \times (1 - \exp(-t/\tau))\tag{8}
$$

where f is the shape of the spheroidal cell [74] and τ is the charging time of the cell membrane, g depends upon the conductivities and R is the radius of the spherical cell. E is the field strength and θ(M) is the angle between normal to the membrane at the position M and direction of the field [55]. The exponential term can be ignored if the pulse length is longer than a few microseconds. Because induction time τ<1µs, the value f is generally 3/2, which is for com‐ pletely insulating membrane [75].
